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Hydration structure of a collagen peptide
Abstract
The collagen triple helix is a unique protein motif defined by the supercoiling of three polypeptide chains in a polyproline II conformation. It is a major domain of all collagen proteins and is also reported to exist in proteins with host defense function and in several membrane proteins. The triple-helical domain has distinctive properties. Collagen requires a high proportion of the post-translationally modified imino acid 4-hydroxyproline and water to stabilize its conformation and assembly. The crystal structure of a collagen-like peptide determined to 1.85 Angstrum showed that these two features may be related.
A detailed analysis of the hydration structure of the collagen-like peptide is presented. The water molecules around the carbonyl and hydroxyprolyl groups show distinctive geometries. There are repetitive patterns of water bridges that link oxygen atoms within a single peptide chain, between different chains and between different triple helices. Overall, the water molecules are organized in a semi-clathrate-like structure that surrounds and interconnects triple helices in the crystal lattice. Hydroxyprolyl groups play a crucial role in the assembly.
The roles of hydroxyproline and hydration are strongly interrelated in the structure of the collagen triple helix. The specific, repetitive water bridges observed in this structure buttress the triple-helical conformation. The extensively ordered hydration structure offers a good model for the interpretation of the experimental results on collagen stability and assembly.
... Unlike an earlier NMR study suggesting that an ordered "water wire" exists parallel to the collagen main axis [20], other ordered arrangements of water were revealed by peptide diffraction studies [21,22]: ...
... There are 28 known collagen families, among which collagens I, II, III, V, and XI are dominant and found in bones, tendons, cartilage, skin, blood vessels, and more [5]. As mentioned above, each collagen polypeptide chain has Water bridges have fascinated the collagen X-ray community because they were envisioned as stabilizing the collagen structure [6,[22][23][24]. These chains were "immobilized relative to bulk water and aligned in the plane transverse to the axis of the molecule", thus explaining the "tendon magic angle effect" seen in magnetic resonance imaging [25]. ...
... In the [(Pro(X)-Pro(Y)-Gly) 10 ] 3 collagen tripeptide model chain, hydrogen bonds between the main chains are responsible for the triple-helix stability. Early fiber diffraction models already deduced the correct interstrand Gly-NH· · · OC-Pro(X) connectivity, where the NH group of the Gly residue acts as a HB donor, and the CO group of the Pro residue in the X position of an adjacent strand acts as an acceptor [14,22]. Therefore there is exactly one such HB per triplet, as depicted schematically in Figure 4. ...
Collagen is a triple-helical protein unique to the extracellular matrix, conferring rigidity and stability to tissues such as bone and tendon. For the [(PPG)10]3 collagen-mimetic peptide at room temperature, our molecular dynamics simulations show that these properties result in a remarkably ordered first hydration layer of water molecules hydrogen bonded to the backbone carbonyl (bb-CO) oxygen atoms. This originates from the following observations. The radius of gyration attests that the PPG triplets are organized along a straight line, so that all triplets (excepting the ends) are equivalent. The solvent-accessible surface area (SASA) for the bb-CO oxygens shows a repetitive regularity for every triplet. This leads to water occupancy of the bb-CO sites following a similar regularity. In the crystal-phase X-ray data, as well as in our 100 K simulations, we observe a 0-2-1 water occupancy in the P-P-G triplet. Surprisingly, a similar (0-1.7-1) regularity is maintained in the liquid phase, in spite of the sub-nsec water exchange rates, because the bb-CO sites rarely remain vacant. The manifested ordered first-shell water molecules are expected to produce a cylindrical electrostatic potential around the peptide, to be investigated in future work.
... Unlike an earlier NMR study suggesting that an ordered "water wire" exists parallel to the collagen main axis [19], other ordered arrangements of water were revealed by the peptide diffraction studies [20,21]: ...
... Water bridges fascinated the collagen X-ray community, because they were envisioned as stabilizing the collagen structure [5,[21][22][23]. These chains were "immobilized relative to bulk water and aligned in the plane transverse to the axis of the molecule", thus explaining the "tendon magic angle effect" seen in magnetic resonance imaging [24]. ...
... In the [(Pro(X)-Pro(Y)-Gly) 10 ] 3 collagen tripeptide model chain, hydrogen bonds between the main chains are responsible for the triple-helix stability. Early fibre diffraction models already deduced the correct interstrand Gly-NH···OC-Pro(X) connectivity, where the NH group of the Gly residue acts as a HB donor, and the CO group of the residue in the X position of an adjacent strand acts as an acceptor [13,21]. Therefore there is exactly one such HB per tripeptide, as depicted schematically in Figure 4. ...
Collagen is a triple-helical protein unique to the extracellular matrix, conferring rigidity and stability to tissues such as bone and tendon. For the [(PPG)10]3 collagen-mimetic peptide at room temperature, our molecular dynamics simulations show that these properties result in a remarkably ordered first hydration layer, of water molecules hydrogen-bonded to the backbone-carbonyl (bb-CO) oxygen atoms. This originates from the following observations. The radius of gyration attests that the PPG triplets are organized along a straight line, so that all triplets (excepting the ends) are equivalent. The solvent accessible surface area (SASA) for the bb-CO oxygens shows a repetitive regularity for every triplet. This leads to bb-CO⋯HOH occupancy following a similar regularity, similar also to the crystal-phase 0-2-1 water occupancy in the P-P-G triplet. The regularity is maintained in spite of the sub-nsec water exchange rate, because the bb-CO sites rarely remain vacant. The manifested ordered first-shell water molecules are expected to produce a cylindrical electrostatic potential around the peptide, that might guide cation diffusion in its vicinity.
... Gelatin is obtained through the partial hydrolysis of collagen fibers extracted from skin, cartilage, bones and/or hair of animals [4] and is composed of a mixture of different polymeric structures: α-chains, β-chains (two α-chains) and γ-chains (three α-chains) [4,5]. These α-chains present a polyproline II conformation, which requires a repetitive peptide Glycine-X-Y sequence, with the iminoacids proline (Pro) and hydroxyproline (Hyp) most frequently located in the X and Y position, respectively [6,7]. This particular molecular configuration plays an important role in the stability of the helical structures associated with the partial renaturation of gelatin [6]. ...
... These α-chains present a polyproline II conformation, which requires a repetitive peptide Glycine-X-Y sequence, with the iminoacids proline (Pro) and hydroxyproline (Hyp) most frequently located in the X and Y position, respectively [6,7]. This particular molecular configuration plays an important role in the stability of the helical structures associated with the partial renaturation of gelatin [6]. Gelatin extraction protocols include the use of acid or alkaline chemicals for hydrolysis at temperatures between 50 and 80 • C [8]. ...
... [η] = 8.6 × 10 −5 ·Mw 0.74 (6) Measurements were performed as previously described [55]. Briefly, concentrations from 2 to 6 g/L of each sample were prepared in 0.1 M NaCl and were left overnight at 4 • C. The flow time of each concentration was determined by measuring the time required for the suspension to flow from the top to the bottom mark of the viscometer (size 50, Z275271-1EA, Sigma Aldrich, St. Louis, MO, USA). ...
For biomedical applications, gelatin is usually modified with methacryloyl groups to obtain gelatin methacryloyl (GelMA), which can be crosslinked by a radical reaction induced by low wavelength light to form mechanically stable hydrogels. The potential of GelMA hydrogels for tissue engineering has been well established, however, one of the main disadvantages of mammalian-origin gelatins is that their sol-gel transitions are close to room temperature, resulting in significant variations in viscosity that can be a problem for biofabrication applications. For these applications, cold-water fish-derived gelatins, such as salmon gelatin, are a good alternative due to their lower viscosity, viscoelastic and mechanical properties, as well as lower sol-gel transition temperatures, when compared with mammalian gelatins. However, information regarding GelMA (with special focus on salmon GelMA as a model for cold-water species) molecular conformation and the effect of pH prior to crosslinking, which is key for fabrication purposes since it will determine final hydrogel’s structure, remains scarce. The aim of this work is to characterize salmon gelatin (SGel) and salmon methacryloyl gelatin (SGelMA) molecular configuration at two different acidic pHs (3.6 and 4.8) and to compare them to commercial porcine gelatin (PGel) and methacryloyl porcine gelatin (PGelMA), usually used for biomedical applications. Specifically, we evaluated gelatin and GelMA samples’ molecular weight, isoelectric point (IEP), their molecular configuration by circular dichroism (CD), and determined their rheological and thermophysical properties. Results showed that functionalization affected gelatin molecular weight and IEP. Additionally, functionalization and pH affected gelatin molecular structure and rheological and thermal properties. Interestingly, the SGel and SGelMA molecular structure was more sensitive to pH changes, showing differences in gelation temperatures and triple helix formation than PGelMA. This work suggests that SGelMA presents high tunability as a biomaterial for biofabrication, highlighting the importance of a proper GelMA molecular configuration characterization prior to hydrogel fabrication.
... Collagen's triple helix features a distinctive helical repetition of around 2.9 nm, with roughly 10 amino acid residues each turn. Additionally, the molecules of collagen supercoil, creating a recognizable banding pattern known as the "Dperiodicity" (Bella, Brodsky and Berman, 1995). Collagen's distinctive stability is attributed to hydroxyproline, a modified form of proline. ...
... Collagen peptides' secondary structures and helical composition may be examined using circular dichroism (CD) spectroscopy, revealing information about their structural domains (Ricard-Blum, 2011). The three-dimensional arrangement of collagen domains has been clarified using sophisticated structural methods including X-ray crystallography, NMR spectroscopy, and electron microscopy (Bella, Brodsky and Berman, 1995). Quality of collagen is essential for bio compatibility and reducing the danger of negative responses when working with living tissues in biomedical and therapeutic applications including tissue engineering and wound healing (Rezvani Ghomi et al., 2021). ...
Particularly in the form of collagen peptides, collagen, an important structural protein present in many animal tissues, has drawn much interest for its possible health advantages. In this study, we look at the methods for isolating, characterizing, and extracting collagen peptides from fish sources. Due to its great availability and sustainability, fish collagen is a possible replacement for conventional collagen sources. Fish collagen peptides must be extracted using a number of processes, including demineralization, deproteinization, and acid or enzymatic treatment. These procedures are necessary to produce collagen that is very pure and bioactive. For characterizing isolated collagen peptides, methods including SDS-PAGE, FTIR spectroscopy, and amino acid analysis are frequently used to evaluate their quality and make-up. The subject of contemporary research has been the biological functions of collagen peptides generated from fish. These peptides have demonstrated a range of health-promoting qualities, including as antioxidant, anti-inflammatory, and anti-aging activities. They have also shown promise in boosting joint health, skin health, and wound healing. Fish collagen peptides are useful additives in medications, cosmetics, and nutritional supplements due to their bioavailability and bioactivity. Additionally, new research has demonstrated the potential of fish collagen peptides to improve gastrointestinal, bone, and cardiovascular health. These results indicate that fish collagen peptides have several uses in the nutraceutical and functional food industries
... The strength and density of the interactions between the tannin molecules and collagen determine leather's hydrothermal stability [35][36][37][38][39], which is generally assessed by measuring the shrinkage temperature using the standard test UNI EN ISO 3380: 2015. However, temperature is an intensive parameter, which cannot explain the mechanism of the shrinkage temperature increasing. ...
... It is therefore very likely that MIDA DD@β-CD poly [2]pseudorotaxanes interact with collagen through multiple hydrogen bonding due to its hydrogen bond donor groups, i.e., phenol and alcohol groups of MIDA DD and β-CD, respectively. Because of their flexibility, the phenolic hydroxyl of PSCCs' side chains are more likely to interact with the side chains of collagen polypeptide chains.During these linking phase, poly [2]pseudorotaxanes displace the interfibrillar water molecules [37,39] while the bulky cyclodextrins threaded along the sidechain of MIDA DD are wrapped into the interfibrillar space. Since water fills the inter-fibrillar spaces and bulges them, expanding the lattice, the accommodation of the PSCC guest-host complex seems to reduce the ability of collagen to be inflated by the action of water. ...
... The compositions of all the studied collagenous materials before and after irradiation were analysed via a number of analytical methods. The determination of the free water and the interstitial water (directly bound to the triple-helix) [28] was performed according to the ISO 6496:1983 standard (Animal feedstuffs -Determination of the moisture content), i. e. via drying up to 160 ± 2 • C for 4 h. ...
... The total water contentsfree and interstitial (directly bound to the triple-helix) [28] including also in the freeze-dried state as determined by the weight loss are shown in Fig. 2. A statistically significantly lower total water content was determined for the materials irradiated in the lyophilized state regardless of the radiation dose than that of the original collagenous materials (Fig. 2A). Conversely, the collagenous materials irradiated in the gel state did not evince statistically significant differences from the original state (Fig. 2B). ...
... Shortly thereafter, Bella and Berman described the precise stereochemistry of Cα-H. . .O=C interactions in collagen, based on their 1.85 Å resolution structure [108] (Figure 3). The stereochemistry of these bonds again aligns perfectly with the one proposed by Huggins, although the authors also appear to have been unaware of Huggins's publication. ...
... This was reaffirmed soon afterward by other authors using a different set of crystal structures, and it was additionally noted, not unexpectedly, that Gly-recurrent in β-sheets-is not a donor in these interactions [107]. Shortly thereafter, Bella and Berman described the precise stereochemistry of Cα-H…O=C interactions in collagen, based on their 1.85 Å resolution structure [108] ( Figure 3). The stereochemistry of these bonds again aligns perfectly with the one proposed by Huggins, although the authors also appear to have been unaware of Huggins's publication. ...
Hydrogen bonds constitute a unique type of non-covalent interaction, with a critical role in biology. Until fairly recently, the canonical view held that these bonds occur between electronegative atoms, typically O and N, and that they are mostly electrostatic in nature. However, it is now understood that polarized C-H groups may also act as hydrogen bond donors in many systems, including biological macromolecules. First recognized from physical chemistry studies, C-H…X bonds were visualized with X-ray crystallography sixty years ago, although their true significance has only been recognized in the last few decades. This review traces the origins of the field and describes the occurrence and significance of the most important C-H…O bonds in proteins and nucleic acids.
... For decades the most credited tanning theory explained the stabilization of the collagen with the formation of cross-links within its triple helix structure [8]. A modification of the supramolecular water sheath also plays a key role in this transformation [9]. Therefore, tannery effluent carries heavy pollution load with different types of tanning materials, salts and large quantities of putrefying suspended matters [10]. ...
... Feed crop residue and feed crop by product production and yield data has been taken from BBS, 2017 [20], for this study considered animals that are cattle and goats. The dressing percentage (DP) and product fraction (P f ) of the hides are calculated using Equations (9) and (10) [11]: DP = Chilled carcass weight Live weight during slaughtering × 100 (9) ...
Leather processing industries consume high volumes of water and chemicals and release effluents into the environment that pollute the surface water and may cause harm to human health. Leather processing involves different wet processing stages such as soaking, liming, chrome tanning, rechroming, neutralization, fatliqouring and dyeing. The pollution generated from the leather processing stages varies in volume, nature and concentrations. Qualitative and quantitative assessments of effluents generated from different stages of leather processing can be useful to understand the stagewise and overall water pollution of leather wet processing and to design and plan pollution abatement initiatives. Water footprints (WF) can help in understanding the total water consumption and water pollution caused by the leather sector. The objectives of this research are to assess the characteristics of effluents generated from different stages of leather processing, calculate the water footprint (WF) and analyze the pollution load of the Bangladesh leather sector. To perform experimental analyses, effluent samples were collected from the following leather processing stages: soaking, liming, deliming and bating, pickling and tanning, wet back, rechroming, neutralization, retanning, dyeing and fatliqouring from four leather processing factories. The key pollution indicating parameters, such as pH, chemical oxygen demand (COD), biological oxygen demand (BOD), total dissolved solid (TDS) and total suspended solid (TSS) of the effluent samples were analyzed. The experimental study showed that almost 52% effluents generate from beam house and tan yard operations, and about 48% effluents generate from post tanning operations. Due to the presence of high amounts of salt, insecticides and bactericides, the effluent generated from the soaking stage contains high BOD and TDS. On the other hand, effluent generated from liming contains the highest amounts of BOD, COD, TDS, and TSS. The reduction or segregation of soaking and liming effluents will be effective in improving the environmental performance of the wet processing of leather. To assess the total water footprint of the leather sector, the water footprint of feed crops and raw hides were calculated, along with the water footprint of the leather processing stages. The water footprints of bovine and ovine crust leather were found to be 34,000 m3/ton and 17,300 m3/ton, respectively. The blue water footprint is higher in soaking, liming and finishing. The green water footprint of leather is mainly contributed by feed crops of farming animals. The grey water footprint was found higher in the soaking, liming, fatliqouring and dyeing stages. About 97% of the water footprints of tanneries are contributed by the wet processing stages. The grey water footprint is the most significant part of the total water footprint of the leather sector, which indicates the impact of high water pollution by the leather processing stages. This study can help to understand the overall scenario of water consumption and water pollution caused by the leather sector in Bangladesh. This study can also be useful in designing sustainable leather products by reducing the total water footprint per unit of leather goods. The systematic approach of this study could be useful for other countries in leather processing.
... The triple helix is stabilised by inter-and intramolecular hydrogen bonding between chains. Bella et al. (1995) reported that multiple hydrogen bonding patterns stabilise the triple helix. These include, i) direct hydrogen bonding among the peptides (i.e., the NH group in glycine in each polypeptide chain forms H-bonds with adjacent peptide CO groups of the other chains), ii) water-mediated hydrogen bonding linking carbonyl groups, and iii) watermediated hydrogen bonding, which links hydroxyproline OH groups and carbonyl groups. ...
... Collagen-starch films show the lowest enthalpy values, and this correlates with reduced crystallinity. It is widely accepted that collagen structure is stabilised by direct and water-mediated hydrogen bonds located within the triple helices (Bella et al., 1995;Brodsky and Ramshaw, 1997;Zhang et al., 2007). ...
Collagen film (casings) obtained from acid-swollen collagen fibres is widely used as an alternative to natural casings for sausage production. However, collagen casings possess weak properties such as low mechanical properties (tensile strength and stiffness) and thermal stability compared to natural casings. Therefore, there is a need industrially to improve these properties. The main purpose of this work was to study the effects of polysaccharides on the properties of acid-swollen collagen pastes and films as a function of collagen paste concentrations (2.5 %, 3.5 % and 4 %wt/wt). In this work, polysaccharides dispersions: cellulose fibres of different length and waxy (WS) and high amylose (HAS) maize starch granules and molecular solutions: Hydroxypropylmethylcellulose (HPMC), Methylcellulose (MC), high molecular weight (GH), low molecular weight guar gum (GM) and Carboxymethylcellulose (CMC) were blended with acid-swollen collagen paste to fabricate collagen films with improved properties such as mechanical properties (tensile strength, stiffness and flexibility) and thermal stability. The viscoelastic of the blend pastes and denaturation of collagen was studied by rheological and thermal techniques. The pure and composite films were studied by sorption, mechanical, spectroscopic, structural, and thermal techniques. The focus of the first part of this study is to investigate the effect of uncharged and negatively charged molecular solutions at comparable low-shear viscosity on the viscoelastic and thermal properties of acid-swollen collagen paste. Dynamic rheological data indicated that the addition of non-charged hydrocolloids: HPMC, MC, GH and GM increased the storage modulus (G’) and loss modulus (G’’) of the acid-swollen collagen paste. By contrast, negatively charged CMC decreased the G’ and G” of the collagen pastes. At the level of addition of non-charged solutions (HPMC, MC, GH and GM) considered in this study, the denaturation temperature of collagen as determined by DSC was not affected while negatively charged CMC increased the denaturation temperature. Composite films containing blends of collagen paste with the individual molecular solutions were formed. Films were characterised for their mechanical, thermal, sorption and structural properties. Collagen/CMC films were not tested due to the difficulty in analysing the films. The thickness of the films increased and was dependent on the collagen concentration as well as the hydrocolloid concentration in the film network. Mechanical data revealed that the addition of hydrocolloids increased the tensile strength (TS), stiffness (YM), and elongation at break (EAB) of the films. Derivatised cellulose showed higher enhancement than the guars. Consistent with the mechanical data, DSC revealed an increase in peak temperature and a decrease in enthalpy of the films with the addition of the polymers. An increase in TS, YM, and EAB and an increase in peak temperatures were dependent on the collagen concentration. XRD data of the composite film showed a reduction in the intensity of the crystalline peak of collagen. FTIR spectra of the films helped to understand the structural changes and the interaction between the collagen and hydrocolloids. The thermal degradation temperature of collagen was not affected, as evidenced by the TGA curves. Furthermore, the composite films showed lower moisture uptake than the pure collagen films. The next study focused on investigating the effect of polysaccharide dispersions, cellulose with different fibre length, waxy and high amylose maize starches at comparable dispersed phase volume on the rheological and thermal properties of acid-swollen collagen paste. The dynamic rheological measurement revealed the dominant elastic behaviour (G’ > G’’) of the blend and control pastes. Cellulose fibres, waxy and high amylose starch granules increased the storage and loss modulus, and values increased with increasing collagen content. The starches exhibited a higher value due to the high concentration used. According to the DSC data, the denaturation of collagen and enthalpy of melting was not affected by the addition of the dispersions. On the other hand, on reheating the blend pastes, the starches lowered the enthalpy of the denatured collagen. Films were made from the blend pastes and were characterised for their mechanical, thermal, sorption and structural properties. The surface of composite films appeared rough because of the protrusion of the cellulose fibres and starch granules. The thickness of the films increased with the addition of the cellulose fibres and starch granules. Values increased with increasing levels of collagen and dispersions concentration in the film-forming paste. Reinforcing collagen films with cellulose fibres increased the mechanical properties (TS, YM and EAB) of the films. The mechanical properties of collagen with starch granules films could not be tested due to the brittleness of the films. DSC data showed that cellulose fibres increased the peak temperature of the films. By contrast, starch granules decreased the peak temperature. The enthalpy of the films was significantly reduced with the addition of cellulose fibres and starch granules. Collagen with starch granules films had the lowest enthalpy values. XRD data showed a decrease in the intensity of the crystalline peak of collagen in the blend films. The thermal stability of collagen was reduced, as evidenced by the TGA data. Additionally, the water uptake of the films decreased with the addition of cellulose fibres and starch granules. For the final study, the effect of collagen pastes (2.5%, 3.5% and 4 % wt) on the pasting properties of waxy (WS), high amylose (HAS) and normal (NS) maize starches were studied using Rapid Viscous Analyser (RVA) at conventional (up to 95 °C) and high-temperature (up to 140 °C) heating modes. Results showed that collagen pastes modified the pasting properties of the starches. At conventional heating mode, high amylose did not show a noticeable pasting profile. The pasting temperature of waxy starch was unaffected by the addition of collagen paste. By contrast, the addition of collagen paste lowered the pasting temperature of normal starches. The viscosities (peak, setback, and breakdown) of NS and WS increased. The final viscosity of WS decreased while that of NS increased with the increase in collagen paste concentration. When the samples are heated to temperatures 140 °C higher, HAS showed a noticeable pasting profile. The pasting temperature of HAS decreased with increasing levels of collagen paste addition. Peak and breakdown viscosities of NS, WS, and HAS increased with increasing collagen paste levels. In contrast, setback and final viscosities reduced.
... Fully assembled collagen molecules are trimers of three identical or different polypeptides referred to as α-chains. A triple-helical domain is generated when each constituent α-chain forms a lefthanded helix which in turn wraps around two other α-chains to form a right-handed super-helix ( Figure 1A) (Ramachandran and Kartha, 1954;Bella et al., 1994;Bella et al., 1995;Okuyama, 2008). To adopt this tertiary structure, the primary structure of the triple helical domain is limited to a three residue repeat glycine-X-Y where X and Y are frequently proline and hydroxyproline residues respectively (Ramachandran and Kartha, 1954;Bella et al., 1994). ...
Collagen VI and collagen XII are structurally complex collagens of the extracellular matrix (ECM). Like all collagens, type VI and XII both possess triple-helical components that facilitate participation in the ECM network, but collagen VI and XII are distinct from the more abundant fibrillar collagens in that they also possess arrays of structurally globular modules with the capacity to propagate signaling to attached cells. Cell attachment to collagen VI and XII is known to regulate protective, proliferative or developmental processes through a variety of mechanisms, but a growing body of genetic and biochemical evidence suggests that at least some of these phenomena may be potentiated through mechanisms that require coordinated interaction between the two collagens. For example, genetic studies in humans have identified forms of myopathic Ehlers-Danlos syndrome with overlapping phenotypes that result from mutations in either collagen VI or XII, and biochemical and cell-based studies have identified accessory molecules that could form bridging interactions between the two collagens. However, the demonstration of a direct or ternary structural interaction between collagen VI or XII has not yet been reported. This Hypothesis and Theory review article examines the evidence that supports the existence of a functional complex between type VI and XII collagen in the ECM and discusses potential biological implications.
... During the self-assembly process, water interacts with collagen, thereby influencing the mechanical properties of the final network. Water surrounds the collagen, and tightly binds to it creating a well-ordered hydration shell (or hydration layer) (4,5) that controls collagen properties. Water is believed to contribute to the structure and stability of the triple helix via the formation of water bridges (6)(7)(8)(9)(10), and it mediates collagen inter-and intra-molecular interactions (11,12). ...
Water is known to play an important role in collagen self-assembly, but it is still largely unclear how water–collagen interactions influence the assembly process and determine the fibril network properties. Here, we use the H 2 O/D 2 O isotope effect on the hydrogen-bond strength in water to investigate the role of hydration in collagen self-assembly. We dissolve collagen in H 2 O and D 2 O and compare the growth kinetics and the structure of the collagen assemblies formed in these water isotopomers. Surprisingly, collagen assembly occurs ten times faster in D 2 O than in H 2 O, and collagen in D 2 O self-assembles into much thinner fibrils, that form a more inhomogeneous and softer network, with a fourfold reduction in elastic modulus when compared to H 2 O. Combining spectroscopic measurements with atomistic simulations, we show that collagen in D 2 O is less hydrated than in H 2 O. This partial dehydration lowers the enthalpic penalty for water removal and reorganization at the collagen–water interface, increasing the self-assembly rate and the number of nucleation centers, leading to thinner fibrils and a softer network. Coarse-grained simulations show that the acceleration in the initial nucleation rate can be reproduced by the enhancement of electrostatic interactions. These results show that water acts as a mediator between collagen monomers, by modulating their interactions so as to optimize the assembly process and, thus, the final network properties. We believe that isotopically modulating the hydration of proteins can be a valuable method to investigate the role of water in protein structural dynamics and protein self-assembly.
... The presence of the recurring tripeptide motif -Gly-X-Y-(where X is Pro and Y is usually hydroxyl proline) in the collagen sequence, alongside a glycine residue at every third position, provides favorable conditions for triple helix formation. The stability of this triple-helical structure significantly relies on the inter-strand assembly mediated by non-covalent interactions, including hydrogen bonding, hydrophobic interactions, ionic interactions, and salt bridges [5][6][7]. The distinctive features of collagen, primarily attributed to its unique triple helical structure and its subsequent fibrillar morphology upon selfassembly, underlie its diverse functional properties. ...
... The role of water on the structure of collagen has been widely investigated in literature using a variety of techniques [21][22][23][24][25]. Such studies have shown hydration can greatly impact the physical properties of collagen [26]. ...
The functional role of collagen piezoelectricity has been under debate since the discovery of piezoelectricity in bone in 1957. The possibility that piezoelectricity plays a role in bone remodeling has generated interest in the investigation of this effect in relevant physiological conditions; however, there are conflicting reports as to whether collagen is piezoelectric in a humid environment. In macroscale measurements, the piezoelectricity in hydrated tendon has been shown to be insignificant compared to dehydrated tendon, whereas, at the nanoscale, the piezoelectric effect has been observed in both dry and wet bone using piezoresponse force microscopy (PFM). In this work, the electromechanical properties of type I collagen from a rat tail tendon have been investigated at the nanoscale as a function of humidity using lateral PFM (LPFM) for the first time. The relative humidity (RH) was varied from 10% to 70%, allowing the piezoelectric behavior to be studied dry, humid, as well as in the hydrated range for collagen in physiological bone (12% moisture content, corresponding to 40–50% RH). The results show that collagen piezoresponse can be measured across the humidity range studied, suggesting that piezoelectricity remains a property of collagen at a biologically relevant humidity.
... Helicoidal portions of collagen molecules are side by side with non-helicoidal parts that are placed at both C and N termini and comprise 16-26 amino acidic residues [4]. The molecular stabilization is guaranteed by hydrogen bond formation within the molecular backbone and between polar amino acid residues and water molecules that surround the triple helix forming a hydration cylinder [5]. Two different models were proposed to describe the helical pattern of the molecular structure of collagen. ...
Type I collagen physiological scaffold for tissue regeneration is considered one of the widely used biomaterials for tissue engineering and medical applications. It is hierarchically organized: five laterally staggered molecules are packed within fibrils, arranged into fascicles and bundles. The structural organization is correlated to the direction and intensity of the forces which can be loaded onto the tissue. For a tissue-specific regeneration, the required macro-and microstructure of a suitable biomaterial has been largely investigated. Conversely, the function of multiscale structural integrity has been much less explored but is crucial for scaffold design and application. In this work, collagen was extracted from different animal sources with protocols that alter its structure. Collagen of tendon shreds excised from cattle, horse, sheep and pig was structurally investigated by wide-and small-angle X-ray scattering techniques, at both molecular and supramolecular scales, and thermo-mechanically with thermal and load-bearing tests. Tendons were selected because of their resistance to chemical degradation and mechanical stresses. The multiscale structural integrity of tendons' collagen was studied in relation to the animal source, anatomic location and source for collagen extraction.
... The determination of the free water and the interstitial water (directly bound to the triple-helix) [27] was performed using the ISO 6496:1983 standard (Animal feedstuffs-Determination of the moisture content), that is, drying to 160 ± 2 °C for 4 h (dryer-Memmert GmbH + Co. KG, Büchenbach, Germany); scales-Mettler-Toledo Ltd. (Prague, Czech Republic). ...
The physical properties and structure of collagen treated with high-pressure technologies have not yet been investigated in detail. The main goal of this work was to determine whether this modern gentle technology significantly changes the properties of collagen. High pressure in the range of 0–400 MPa was used, and the rheological, mechanical, thermal, and structural properties of collagen were measured. The rheological properties measured in the area of linear viscoelasticity do not statistically significantly change due to the influence of pressure or the duration of pressure exposure. In addition, the mechanical properties measured by compression between two plates are not statistically significantly influenced by pressure value or pressure hold time. The thermal properties Ton and ΔH measured by differential calorimetry depend on pressure value and pressure hold time. Results from amino acids and FTIR analyses show that exposure of collagenous gels to high pressure (400 MPa), regardless of applied time (5 and 10 min), caused only minor changes in the primary and secondary structure and preserved collagenous polymeric integrity. SEM analysis did not show changes in collagen fibril ordering orientation over longer distances after applying 400 MPa of pressure for 10 min.
... Moreover, the COL/PGT/PES membrane had a higher oxygen content than the nascent PES membrane. The increase in oxygen content is mainly due to the rich oxygen content of COL [40]. ...
This study focuses on the fabrication of low-fouling polyethersulfone ultrafiltration (PES UF) membranes using collagen (COL) and polyphenon 60 from green tea (PGT) as additives. The casting solution was prepared by dissolving COL and PGT in PES solution with N-methyl-2-pyrrolidone as solvent. A series of characterizations and performance examinations, including scanning electron microscopy–energy-dispersive X-ray spectroscopy (SEM-EDX) mapping, Fourier transform infrared spectroscopy (FTIR), contact angle (CA) measurement, pure water flux (PWF), porosity, antifouling test using bovine serum albumin (BSA) and humic acid (HA) solution, antibacterial test using E. coli and S. mutans, were conducted. The addition of COL and PGT not only significantly increased PWF, water uptake, porosity, and hydrophilicity but also increased solute rejection and flux recovery. The membrane prepared with the addition of COL (1 wt%) and PGT (3 wt%) demonstrated PWF of 353.9 L/m2. h, water uptake of 238.8%, porosity of 75.1%, CA of 36.9°, and solute rejection of 98.5%. Further, the flux recovery reached 85.08% and 78.78% after anti fouling examination using BSA and HA solutions, respectively. The COL/PGT/PES membrane also demonstrated superior dynamic antibacterial filtration with 93.5% bacteria removal of E. coli and highly efficient antibacterial activity above ∼98% against both E. coli and S. mutans. Thus, the COL/PGT/PES membrane exhibited significant antifouling and antibacterial properties and better performance than the nascent PES UF membrane.
... The exact compositions of the two collagenous gels were analysed by means of a number of analytical methods. The determination of the free water (eliminated via lyophilization and drying at 105 • C) and the interstitial water -directly bound to the triple-helix (Bella et al., 1995) was performed according to the ISO 6496:1983 standard (Animal feedstuffs -Determination of the moisture content), i.e. via drying to 103 ± 2 • C for 4 h and drying to 160 ± 2 • C for 3 h, respectively (N = 5). The collagen gels were frozen at − 30 • C and lyophilized (BenchTop 4KZL, VirTis) for further analysis. ...
... The figure is adapted with permission from Andriotis et al.[36] . (B) The tiple helix of a 30-residue long part of a collagen molecule (PDB: 1CGD) is shown on top without and below with the water monolayer as predicted by[95] , with permission under a CC By 4.0 license from the RCSB PDB (10.2210/pdb1CGD/pdb). ...
Collagen fibrils are the fundamental structural elements in vertebrate animals and compose a framework that provides mechanical support to load-bearing tissues. Understanding how these fibrils initially form and mechanically function has been the focus of a myriad of detailed investigations over the last few decades. From these studies a great amount of knowledge has been acquired as well as a number of new questions to consider. In this review, we examine the current state of our knowledge of the mechanical properties of extant fibrils. We emphasize on the mechanical response and related deformation of collagen fibrils upon tension, which is the predominant load imposed in most collagen-rich tissues. We also illuminate the gaps in knowledge originating from the intriguing results that the field is still trying to interpret.
Statement of significance
: Collagen is the result of millions of years of biological evolution and is a unique family of proteins, the majority of which provide mechanical support to biological tissues. Cells produce collagen molecules that self-assemble into larger structures, known as collagen fibrils. As simple as they appear under an optical microscope, collagen fibrils display a complex ultrastructural architecture tuned to the external forces that are imposed upon them. Even more complex is the way collagen fibrils deform under loading, and the nature of the mechanisms that drive their formation in the first place. Here, we present a cogent synthesis of the state-of-knowledge of collagen fibril mechanics. We focus on the information we have from in vitro experiments on individual, isolated from tissues, collagen fibrils and the knowledge available from in silico tests.
... Of principal interest to this Perspective is the subfamily of fibrillar collagens, whose coiledcoil molecules form yet another higher-level superstructure: through a combination of covalent cross-linking and non-covalent interactions, they are assembled into fibrils of a diameter 50-100 nm. 3,[7][8][9] The fibrils are assembled into larger fibres and can further form a cross-linked network that serves as the structural scaffold of the extracellular matrix (ECM) in many biological tissues. ...
It is well-known that collagen is the most abundant protein in the human body; however, what is not often appreciated is its fascinating physical chemistry and molecular physics. In this Perspective, we aim to expose some of the physicochemical phenomena associated with the hydration of collagen and to examine the role collagen's hydration water plays in determining its biological function as well as applications ranging from radiology to bioengineering. The main focus is on the Magic-Angle Effect, a phenomenon observed in Nuclear Magnetic Resonance (NMR) spectroscopy and Magnetic Resonance Imaging (MRI) of anisotropic collagenous tissues such as articular cartilage and tendon. While the effect has been known in NMR and MRI for decades, its exact molecular mechanism remains a topic of debate and continuing research in scientific literature. We survey some of the latest research aiming to develop a comprehensive molecular-level model of the Magic-Angle Effect. We also touch on other fields where understanding of collagen hydration is important, particularly nanomechanics and mechanobiology, biomaterials, and piezoelectric sensors.
... Hydroxyproline moieties on collagen can also play both roles. 90,91 It is expected to exhibit donating properties when involved in interactions with basic centers, such as carboxylate oxygen atoms. This was confirmed by MD calculations. ...
Hyaluronan - collagen composites have found numerous biomedical applications. Understanding the interactions between hyaluronan and collagen is particularly important in the context of joint cartilage function and the treatment of joint- diseases. There are many factors that affect the affinity of collagen for hyaluronan. One of the important features is the ratio of 3- or 4-hydroxy proline to proline residues. This article presents results from molecular dynamics calculations of HA-collagen type II interactions with hyaluronan. The applied protocol employed docking and geometry optimizations of complexes built using collagen structures with different number of hydroxyl groups attached to proline moieties. It was established that the hydroxyproline/proline ratio affects both structural and energetical features of the collagen-hyaluronan complex. Proline hydroxylatinon was found to significantly influence the number of all identified types of molecular forces, namely hydrophobic interactions, ionic contacts, water bridges and hydrogen bonds, which can be formed between collagen and hyaluronan. Furthermore, an increase of the hydroxyproline/proline ratio in the collagen chain increaeses the binding efficiency to hyaluronan. This is illustrated by the linear correlation between binding free energy and the hydroxylation degree. The comparison of the results obtained for 3 and 4 hydroxylation indicates that hydroxyl group attachment position play a minor role in the complex stabilization. However, a slightly stronger affinity was observed for the latter case. In order to evaluate the role of the aqueous environment on the collagen-hyaluronan complex stability, the entalpic and entropic contributions to the free energy of solvatation were analyzed.
... Next, we surveyed existing crystal structures of CMP triplehelices in the Protein Data Bank (PDB, see Table S3 †) to search for evidence of structural differences between CMPs with POG-and GPO-terminal repeats. 23,24,[32][33][34][35][36][37][38][39][40][41] We analyzed the Bfactor of each CMP structure as it oen correlates with the exibility and internal motion in protein crystallography. 42 We plotted normalized B-factors of all non-hydrogen atoms along each CMP triple-helix: while all structures have elevated structural exibility at the termini, a general trend of higher terminal B-factor was noted for the POG-sequences (Fig. 3a, S3 and S4 †). ...
Nearly 30% of human proteins have tandem repeating sequences. Structural understanding of the terminal repeats is well-established for many repeat proteins with the common α-helix and β-sheet foldings. By contrast, the sequence-structure interplay of the terminal repeats of the collagen triple-helix remains to be fully explored. As the most abundant human repeat protein and the most prevalent structural component of the extracellular matrix, collagen features a hallmark triple-helix formed by three supercoiled polypeptide chains of long repeating sequences of the Gly-X-Y triplets. Here, with CD characterization of 28 collagen-mimetic peptides (CMPs) featuring various terminal motifs, as well as DSC measurements, crystal structure analysis, and computational simulations, we show that CMPs only differing in terminal repeat may have distinct end structures and stabilities. We reveal that the cross-chain hydrogen bonding mediated by the terminal repeat is key to maintaining the triple-helix's end structure, and that disruption of it with a single amide to carboxylate substitution can lead to destabilization as drastic as 19 °C. We further demonstrate that the terminal repeat also impacts how strong the CMP strands form hybrid triple-helices with unfolded natural collagen chains in tissue. Our findings provide a spatial profile of hydrogen bonding within the CMP triple-helix, marking a critical guideline for future crystallographic or NMR studies of collagen, and algorithms for predicting triple-helix stability, as well as peptide-based collagen assemblies and materials. This study will also inspire new understanding of the sequence-structure relationship of many other complex structural proteins with repeating sequences.
... In the collagen triple helical structure, the -OH group of hydroxyproline is projected outside and favors inter-chain interactions and molecular staggering to promote self-assembly. 134 The presence of hydroxyproline in the cell receptor interaction site is essential for cell compatibility and viability. The presence of hydroxyproline in CLP protein resulted in a higher proliferation of 3T3 mouse fibroblast cells and human endothelial cells. ...
Collagen occurs in nature with a dedicated triple helix structure and is the most preferred biomaterial in commercialized medical products. Recombinant collagen emerge as sustainable alternate source that overcomes existing demerits.
... The ΔTs showed the order of PFS-50 retanned leather < PFS-63 retanned leather < PFS-74 retanned leather < control leather. Collagen is surrounded by a supramolecular water sheath nucleated at the hydroxyproline side chains [37]. When PFS with lower DS was introduced into collagen, it can replace more water molecules around collagen than the PFS with higher DS (Fig. 4). ...
Effect of retanning on the thermal stability of leather is eliciting increasing attention. However, the relationship between the hydrophilicity of retanning agents and the heat resistance of leather and the corresponding mechanism remain unclear. Herein, phenolic formaldehyde syntans (PFSs) were selected as models to explore the effect of the hydrophilicity of retanning agents on the thermal stability of retanned leather. The thermal stability of leather was closely correlated to the hydrophilic group content (sulfonation degree) of PFSs. As the sulfonation degree increased, the water absorption rate of PFSs and their retanned leathers decreased, whereas the thermal stability of leather increased. Molecular dynamics simulation results proved that the introduction of PFSs could reduce the binding ability of collagen molecules with water and thus decreased the water molecules around the PFS-treated collagen. These results may provide guidance for the tanners to select retanning agents reasonably to improve the thermal stability of leather.
Graphical Abstract
... In this study, type-I collagen, which is the major organic component of demineralized dentin, served as the receptor for molecular docking with MDP., Docking simulations were performed using the AutoDock program package according to the method mentioned above. Three crystal structures of type-I collagen with different amino acid residues were selected from the Protein Data Bank (PDB) database (PDB IDs: 1CGD, 4OY5, and 1QSU) [32][33][34]. ...
Objectives:
10-Methacryloyloxydecyl dihydrogen phosphate (MDP) has been regarded as the most effective dentin-bonding monomer for more than 20 years. Although the dentin-bonding promoting effect of MDP has been well demonstrated, the mechanisms by which it benefits the stably of collagen within the adhesive-dentin hybrid layer are not currently fully understood. The objective of this study was to investigate the roles of MDP and its calcium salt in preserving the adhesive-dentin hybrid layer.
Methods:
MDP-conditioned collagen was investigated by Fourier-transform infrared spectroscopy, Ultraviolet-visible spectroscopy, and molecular docking. The structural changes to the dentin surface upon acid-etching and MDP-conditioning were observed by SEM. X-ray diffraction and nuclear magnetic resonance were used to investigate the chemical interactions between MDP and HAp. The collagen-protecting effects of MDP and its Ca salt were investigated using in-situ zymography, rhMMP-9 colorimetric assay, hydroxyproline assay, and molecular docking.
Results:
MDP forms a stable collagen-phosphate complex through hydrogen bonding with the collagen in dentin. Furthermore, it generates MDP-Ca salts that are deposited on the dentin collagen scaffold, protecting it from degradation. Moreover, both free MDP and the MDP-Ca salt inhibit matrix metallopeptidase and exogenous proteases, with the inhibitory effect of the calcium salt being significantly stronger than that of the free form.
Significance:
MDP-based adhesives preserve the collagen within the hybrid layer by simultaneously improving collagen's resistance to exogenous enzymes and inhibiting MMP activity, both of which contribute to the longevity of dentin-resin bonding.
... This signal is a marker of the distance between two adjacent amino acids along the central axis of the triple helix of collagen. Both of them are related to the crystalline structural component of type I collagen [24][25][26][27]. ...
Type I collagen is the main component of the extracellular matrix that acts as the physical and biochemical support of tissues. Thanks to its characteristics, collagen is widely employed as a biomaterial for implantable device fabrication and as antiaging food supplementation. Because of the BSE transmission in the 1990s, aquatic animals have become a more suitable extraction source than warm-blooded animals. Moreover, as recently demonstrated, a supplementing diet with fish collagen can increase the body’s collagen biosynthesis. In this context, Tilapia feeding was supplemented with hydrolyzed collagen in order to enhance the yield of extracted collagen. Tilapia skin was investigated with wide and small angle scattering techniques, analyzing the collagen structure from the submolecular to the nanoscale and correlated with Differential Scanning Calorimetry (DSC) measurements. Our studies demonstrated that the supplementation appears to have an effect at the nanoscale in which fibrils appear more randomly oriented than in fish fed with no supplemented feed. Conversely, no effect of a collagen-rich diet was observed at the submolecular scale.
... Three typical type I collagens with different amino acid residue sequences (1CGD, 4OY5, and 1QSU) were obtained from the Protein Data Bank as comparative models [23][24][25]. Based on the molecular information provided by PubChem (https://pubchem.ncbi.nlm.nih. ...
Objectives
: To evaluate the interactions of two phosphate ester monomers [10-methacryloyloxydecyl dihydrogen phosphate (10-MDP) and dipentaerythritol penta-acrylate phosphate (PENTA)] with hydroxyapatite and collagen and understand their influence on dentine bonding.
Methods
: Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, X-ray diffraction, nuclear magnetic resonance, ultraviolet-visible, and molecular docking were applied for separately evaluating the interactions of two monomers with hydroxyapatite and collagen. Hydrophilicity tests and morphological observation were employed to characterize pretreated dentine. Microtensile bond strength (μTBS) and nanoleakage were investigated to evaluate the bonding performance. Hydroxyproline assay, in situ zymography, and matrix metalloproteinase-9 (MMP-9) activity assay were used to confirm the MMP inhibition.
Results
: Chemoanalytic characterization confirmed the interactions of 10-MDP and PENTA with hydroxyapatite and collagen. The interactions of PENTA were weaker than 10-MDP. PENTA possessed better dentine tubule sealing after etching than 10-MDP. Dentine treated with PENTA was more hydrophilic than 10-MDP. 10-MDP and PENTA treating significantly increased the initial μTBS than the control group without primer conditioning. μTBS decreased significantly during aging, and the decrease was more severe in the PENTA group than 10-MDP. The 10-MDP and PENTA groups exhibited relatively less fluorescence than the control. The relative inhibition percentages of MMP-9 decreased in the order of 10-MDP–Ca salt, 10-MDP and PENTA. The 10-MDP, PENTA, and 10-MDP–Ca salt groups showed significantly lower hydroxyproline contents than the control.
Conclusions
: Although PENTA adsorbed on hydroxyapatite, it did not form a stable calcium salt. The interactions of 10-MDP with hydroxyapatite and collagen are different than those of PENTA.
Clinical Significance
: The sealing of dentinal tubules by PENTA and the inhibition of MMP by 10-MDP and its calcium salts contribute to improving the dentine bonding durability.
... Particularly at the molecular level, two main age-related structural alterations occur within the TC, namely the change in diameter (due to the change in the degree of hydration of the molecule) and the change in the number of H-bonds Verz and AuthorAnonymous, 1963;Verzár, 1969). It has been suggested that a weakening of water/TC interactions occurs with age, resulting in a loss of hydration of the molecule and a corresponding decrease in its diameter (Bella et al., 1995). Moreover, heat denaturation experiments showed that H-bonds in the collagen molecule decrease with age (Verzár, 1969) confirmed by (Verz and AuthorAnonymous, 1963) who observed a higher frequency of H-bonds in the collagen molecule of young tendons compared with older ages. ...
With osteoporosis and aging, structural changes occur at all hierarchical levels of bone from the molecular scale to the whole tissue, which requires multiscale modeling to analyze the effect of these modifications on the mechanical behavior of bone and its remodeling process. In this paper, a novel hybrid multiscale model for cortical bone incorporating the tropocollagen molecule based on the combination of finite element method and different homogenization techniques was developed. The objective was to investigate the influence of age-related structural alterations that occur at the molecular level, namely the decrease in both molecular diameter (due to the loss of hydration) and number of hydrogen bonds, on mechanical properties of the bone tissue. The proposed multiscale hierarchical approach is divided in two phases: (i) in Step 0, a realistic 3D finite element model for tropocollagen was used to estimate the effective elastic properties at the molecular scale as a function of the collagen molecule’s degree of hydration (represented by its external diameter) and the number of its intramolecular hydrogen bonds, and (ii) in Steps 1–10, the effective elastic constants at the higher scales from mineralized fibril to continuum cortical bone tissue were predicted analytically using homogenization equations. The results obtained in healthy mature cortical bone at different scales are in good agreement with the experimental data and multiscale models reported in the literature. Moreover, our model made it possible to visualize the influence of the two parameters (molecular diameter and number of hydrogen bonds) that represent the main age-related alterations at the molecular scale on the mechanical properties of cortical bone, at its different hierarchical levels. Keywords: Bone aging, multiscale model, tropocollagen, cortical bone, finite element modeling, homogenization method.
... The existence of 4-Hyp is critical for the stability of the tropocollagen, but this is not fully understood. One of the considerations is that collagen is stabilized mainly by water molecules surrounding collagen that provide a formation of a network of hydrogen bonds between the hydroxyl group of 4-Hyp and main-chain's oxygens [33]. Another consideration is that 4-Hyp reduces the number of conformations available to the random coil of triple helix [34]. ...
... The existence of 4-Hyp is critical for the stability of the tropocollagen, but this is not fully understood. One of the considerations is that collagen is stabilized mainly by water molecules surrounding collagen that provide a formation of a network of hydrogen bonds between the hydroxyl group of 4-Hyp and main-chain's oxygens [33]. Another consideration is that 4-Hyp reduces the number of conformations available to the random coil of triple helix [34]. ...
Hydroxyproline is one of the most prevalent amino acids in animal proteins. It is not a genetically encoded amino acid, but, rather, it is produced by the post-translational modification of proline in collagen, and a few other proteins, by prolyl hydroxylase enzymes. Although this post-translational modification occurs in a limited number of proteins, its biological significance cannot be overestimated. Considering that hydroxyproline cannot be re-incorporated into pro-collagen during translation, it should be catabolized following protein degradation. A cascade of reactions leads to production of two deleterious intermediates: glyoxylate and hydrogen peroxide, which need to be immediately converted. As a result, the enzymes involved in hydroxyproline catabolism are located in specific compartments: mitochondria and peroxisomes. The particular distribution of catabolic enzymes in these compartments, in different species, depends on their dietary habits. Disturbances in hydroxyproline catabolism, due to genetic aberrations, may lead to a severe disease (primary hyperoxaluria), which often impairs kidney function. The basis of this condition is accumulation of glyoxylate and its conversion to oxalate. Since calcium oxalate is insoluble, children with this rare inherited disorder suffer from progressive kidney damage. This condition has been nearly incurable until recently, as significant advances in substrate reduction therapy using small interference RNA led to a breakthrough in primary hyperoxaluria type 1 treatment.
... Although collagen has the potential to be rehydrated and retain moisture, this protein likely has lower hygroscopic power than polysaccharides present in Hyalurosmooth ® . This may be because collagen has a more complex chemical structure than galactomannans and hydroxyethyl cellulose have fewer water molecule bindings with the triple helix [20]. It is necessary to assess the hydrolyzed collagen with other concentrations and verify if this protein works as a preservative of Aedes eggs. ...
Dengue and other Aedes-borne diseases have dramatically increased over the last decades. The Sterile Insect Technique (SIT) has been successfully used as part of integrated pest strategies to control populations of insect-plant and livestock pests and is currently being tested as a potential method to reduce mosquito populations in an environmentally friendly approach. However, during the mass rearing steps needed to produce millions of mosquitoes, egg storage and preservation are essential for a certain amount of time. Eggs of Aedes aegypti have a chorionic pad that functions as a sticky substance to glue them onto the inner walls of larval breeding sites. The chorionic pad is chemically made of hyaluronic acid, a hygroscopic compound, responsible to protect them from desiccation over time. Two commercial products with hygroscopic properties, hydrolyzed collagen, and Hyalurosmooth®, both were tested to assess their ability to prolong egg life storage for A. aegypti and A. albopictus. Results showed that 85–95% of Ae. aegypti eggs were able to hatch up to week 8 after being treated with both hydrophilic compounds, compared with the control 66.3%. These two substances showed promising effects for keeping Ae. aegypti eggs viable during prolonged storage in mass rearing insect production focused on vector control SIT programs.
... water removal (Masic et al., 2015). Within the tightly bound water pool, three distinct collagen-associated water compartments have been identified: single and double water bridges (between alpha helices too distant for direct hydrogen bonding), cleft water within the grooves of the triple helix, and water of the interfacial monolayer (Bella et al., 1995;Brodsky and Persikov, 2005;Lazarev et al., 1992;Wilson et al., 2005). Water also determines collagen's efficient functioning at temperatures (Trebacz and Wojtowicz, 2005): dry proteins cannot unfold upon heating or cooling thus the ability to denature due to temperature is likely caused by the presence of bound water in the system. ...
Water constitutes roughly a quarter of the cortical bone by volume yet can greatly influence mechanical properties and tissue quality. There is a growing appreciation for how water can dynamically change due to age, disease, and treatment. A key emerging area related to bone mechanical and tissue properties lies in differentiating the role of water in its four different compartments, including free/pore water, water loosely bound at the collagen/mineral interfaces, water tightly bound within collagen triple helices, and structural water within the mineral. This review summarizes our current knowledge of bone water across the four functional compartments and discusses how alterations in each compartment relate to mechanical changes. It provides an overview on the advent of- and improvements to- imaging and spectroscopic techniques able to probe nano-and molecular scales of bone water. These technical advances have led to an emerging understanding of how bone water changes in various conditions, of which aging, chronic kidney disease, diabetes, osteoporosis, and osteogenesis imperfecta are reviewed. Finally, it summarizes work focused on therapeutically targeting water to improve mechanical properties.
Overview of natural and engineered material-binding peptides and the molecular forces crucial for their (material-specific) binding to material surfaces.
Collagen is the most abundant structural protein in mammals. Type I collagen in its fibril form has a characteristic pattern structure that alternates two regions called gap and overlap. The structure and properties of collagens are highly dependent on the water and mineral content of the environment. Here, we apply 3D AFM to characterize at angstrom-scale resolution the interfacial water structure of collagen nanoribbons. For a neutral tip, the interfacial water structure is characterized by the oscillation of the water particle density distribution with a value of 0.3 nm (hydration layers). The interfacial structure does not depend on the collagen region. For a negatively charged tip, the interfacial structure might depend on the collagen region. Hydration layers are observed in overlap regions, while in gap regions, the interfacial solvent structure is dominated by electrostatic interactions. These interactions generate interlayer distances of 0.2 nm.
Magnesium ions are highly enriched in early stage of biological mineralization of hard tissues. Paradoxically, hydroxyapatite (HAp) crystallization is inhibited significantly by high concentration of magnesium ions. The mechanism to regulate magnesium‐doped biomimetic mineralization of collagen fibrils has never been fully elucidated. Herein, it is revealed that citrate can bioinspire the magnesium‐stabilized mineral precursors to generate magnesium‐doped biomimetic mineralization as follows: Citrate can enhance the electronegativity of collagen fibrils by its absorption to fibrils via hydrogen bonds. Afterward, electronegative collagen fibrils can attract highly concentrated electropositive polyaspartic acid‐Ca&Mg (PAsp‐Ca&Mg) complexes followed by phosphate solution via strong electrostatic attraction. Meanwhile, citrate adsorbed in/on fibrils can eliminate mineralization inhibitory effects of magnesium ions by breaking hydration layer surrounding magnesium ions and thus reduce dehydration energy barrier for rapid fulfillment of biomimetic mineralization. The remineralized demineralized dentin with magnesium‐doped HAp possesses antibacterial ability, and the mineralization mediums possess excellent biocompatibility via cytotoxicity and oral mucosa irritation tests. This strategy shall shed light on cationic ions‐doped biomimetic mineralization with antibacterial ability via modifying collagen fibrils and eliminating mineralization inhibitory effects of some cationic ions, as well as can excite attention to the neglected multiple regulations of small biomolecules, such as citrate, during biomineralization process.
Amphiphilic biomolecules are abundant in mineralization front of biological hard tissues, which play a vital role in osteogenesis and dental hard tissue formation. Amphiphilic biomolecules function as biosurfactants, however, their biosurfactant role in biomineralization process has never been investigated. This study, for the first time, demonstrates that aggregated amorphous calcium phosphate (ACP) nanoparticles can be reversed into dispersed ultrasmall prenucleation clusters (PNCs) via breakdown and dispersion of the ACP nanoparticles by a surfactant. The reduced surface energy of ACP@TPGS and the electrostatic interaction between calcium ions and the pair electrons on oxygen atoms of C‐O‐C of D‐α‐tocopheryl polyethylene glycol succinate (TPGS) provide driving force for breakdown and dispersion of ACP nanoparticles into ultrasmall PNCs which promote in vitro and in vivo biomimetic mineralization. The ACP@TPGS possesses excellent biocompatibility without any irritations to oral mucosa and dental pulp. This study not only introduces surfactant into biomimetic mineralization field, but also excites attention to the neglected biosurfactant role during biomineralization process.
Collagen is one of the most studied proteins due to its fundamental role in creating fibrillar structures and supporting tissues in our bodies. Accordingly, collagen is also one of the most used proteins for making tissue-engineered scaffolds for various types of tissues. To date, the high abundance of hydroxyproline (Hyp) within collagen is commonly ascribed to the structure and stability of collagen. Here, we hypothesize a new role for the presence of Hyp within collagen, which is to support proton transport (PT) across collagen fibrils. For this purpose, we explore here three different collagen-based hydrogels: the first is prepared by the self-assembly of natural collagen fibrils, and the second and third are based on covalently linking between collagen via either a self-coupling method or with an additional cross-linker. Following the formation of the hydrogel, we introduce here a two-step reaction, involving (1) attaching methanesulfonyl to the -OH group of Hyp, followed by (2) removing the methanesulfonyl, thus reverting Hyp to proline (Pro). We explore the PT efficiency at each step of the reaction using electrical measurements and show that adding the methanesulfonyl group vastly enhances PT, while reverting Hyp to Pro significantly reduces PT efficiency (compared with the initial point) with different efficiencies for the various collagen-based hydrogels. The role of Hyp in supporting the PT can assist in our understanding of the physiological roles of collagen. Furthermore, the capacity to modulate conductivity across collagen is very important to the use of collagen in regenerative medicine.
Background:
Dielectric properties of biological tissues are biophysical parameters; in lung they change with amount of air, blood and parenchyma. Remote Dielectric Sensing (ReDS™) technology measures dielectric properties of lung tissues quantifying the content of fluids inside the scan volume. We aimed to evaluate the reliability of ReDS™ measure in Idiopathic Pulmonary Fibrosis (IPF) patients and in healthy volunteers, and to investigate the correlation of ReDS™ score with clinical, radiological and functional parameters.
Methods:
We conducted a prospective observational study, including 52 patients with diagnosis of IPF and 17 healthy volunteers; for each patient we recorded: complete functional evaluation, dyspnoea score (mMRC scale), Usual Interstitial Pneumonia (UIP) Computed Tomography (CT) pattern (UIP definite or probable) and ReDS™ measure (expressed in %).
Results:
ReDS™ measure was reported as correct both in patients and controls, the firsts with higher scores (33.8% vs 29.1%, p = 0.003). In IPF patients we observed a significant inverse correlation with ReDS™ score and Forced Vital Capacity (FVC), Vital Capacity (VC) and Total Lung Capacity (TLC) measures and, when we considered only patients with UIP definite CT pattern, the correlation was inverse with FVC, VC, TLC, DLCO. In IPF patients the higher was mMRC dyspnoea index, the higher was ReDS™ score. No significant correlations were observed between ReDS™ score and functional parameters in healthy controls.
Discussion:
We demonstrated a correlation of ReDS™ scores with some functional (mainly indicative or diagnostic for restriction) and clinical parameters in IPF patients; the score was correlated with density of tissues possibly quantifying tissue fibrosis in IPF patients.
This review presents recent advances regarding biomass-based nanomaterials, focusing on their surface interactions. Plant biomass-based nanoparticles, like nanocellulose and lignin from industry side streams, hold great potential for the development of lightweight, functional, biodegradable, or recyclable material solutions for a sustainable circular bioeconomy. However, to obtain optimal properties of the nanoparticles and materials made thereof, it is crucial to control the interactions both during particle production and in applications. Herein we focus on the current understanding of these interactions. Solvent interactions during particle formation and production, as well as interactions with water, polymers, cells and other components in applications, are addressed. We concentrate on cellulose and lignin nanomaterials and their combination. We demonstrate how the surface chemistry of the nanomaterials affects these interactions and how excellent performance is only achieved when the interactions are controlled. We furthermore introduce suitable methods for probing interactions with nanomaterials, describe their advantages and challenges, and introduce some less commonly used methods and discuss their possible applications to gain a deeper understanding of the interfacial chemistry of biobased nanomaterials. Finally, some gaps in current understanding and interesting emerging research lines are identified.
Collagen is an active macromolecule, but direct comparisons between different sources were scarce. The purpose of this study was to compare collagen from three fish species to find a collagen with higher bioactivity. Collagen from the swim bladder of Ctenopharyngodon idella, Nibea coibor and Protonibea diacanthus were isolated and named CASC, NASC and PASC, respectively. SDS-PAGE and spectroscopic analysis indicated that ASCs contained triple-helix type I collagen. The microstructure of collagen after lyophilization was uniform and porous. In addition, DSC analysis of the ASCs showed that the denaturation temperature (Td) and melting temperature (Tm) were 79 °C–93 °C and ∼216 °C, respectively. Moreover, the antioxidant activity of ASCs and its effect on fibroblast viability and collagen synthesis were investigated. Current studies have shown that ASCs had antioxidant capacity and were not toxic to fibroblasts. NASC had a higher antioxidant capacity (Hydroxyl, DPPH and ABTS radical scavenging rates were 33.73%, 33.49% and 89.81%, respectively) and significantly promoted fibroblast viability compared to the other two ASCs. It was further found that NASC can promote collagen synthesis in fibroblasts. Collagen extracted from swim bladder may be a promising functional product for the food, cosmetics and biomedical fields.
Dental pulp regeneration exploits tissue engineering concepts using stem cells/scaffolds/growth-factors. Extracted collagen is commonly used as a biomaterial-scaffold due to its biocompatibility/biodegradability and mimics the natural extracellular matrix. Adding biomolecules into a collagen-scaffold enhanced pulp regeneration. Acemannan, β-(1-4)-acetylated-polymannose, is a polysaccharide extracted from aloe vera. Acemannan is a regenerative biomaterial. Therefore, acemannan could be a biomolecule in a collagen-scaffold. Here, acemannan and native collagen were obtained and characterized. The AceCol-scaffold's physical properties were investigated using FTIR, SEM, contact angle, swelling, pore size, porosity, compressive modulus, and degradation assays. The AceCol-scaffold's biocompatibility, growth factor secretion, osteogenic protein expression, and calcification were evaluated in vitro. The AceCol-scaffolds demonstrated higher hydrophilicity, swelling, porosity, and larger pore size than the collagen scaffolds (p < 0.05). Better cell-cell and cell-scaffold adhesion, and dentin extracellular matrix protein (BSP/OPN/DSPP) expression were observed in the AceCol-scaffold, however, DSPP expression was not detected in the collagen group. Significantly increased cellular proliferation, VEGF and BMP2 expression, and mineralization were detected in the AceCol-scaffold compared with the collagen-scaffold (p < 0.05). Computer simulation revealed that acemannan's 3D structure changes to bind with collagen. In conclusion, the AceCol-scaffold synergistically provides better physical and biological properties than collagen. The AceCol-scaffold is a promising material for tissue regeneration.
In this study, we aimed to examine the effect of 3-methacryloxypropyltrimethoxysilane (MPS) on dentin collagen and the impact of MPS and 10-methacryloyloxydecyl dihydrogen phosphate (MDP) together and separately on resin-dentin bonding. Eight groups of primers were prepared: control group, MDP, MPS5, MPS5 + MDP, MPS10, MPS10 + MDP, MPS15, and MPS15 + MDP. The potential interaction between MPS and collagen was assessed by molecular dynamics, contact angle measurement, zeta potential measurement, and chemoanalytic characterization using X-ray photoelectron spectroscopy, Raman spectroscopy, Fourier-transform infrared (FTIR) spectroscopy, and ultraviolet-visible spectroscopy. Microtensile bond strength (μTBS) and nanoleakage were evaluated after 24 h or 12 months of water storage. In situ zymography was used to evaluate the enzyme activity at the bonded interface. According to chemoanalytic characterization and molecular dynamics, a weak interaction between MPS and collagen was observed. MPS enhanced the hydrophobicity and negative charge of the collagen surface (P < 0.05). Applying an MDP-containing primer increased μTBS (P > 0.05) and reduced fluorescence after 24 h of water storage. Water storage for 12 months decreased μTBS (P < 0.05) and increased nanoleakage for all groups. MPS conditioning did not change μTBS and nanoleakage after 24 h of water storage or aging. The MPS10 + MDP and MPS15 + MDP groups presented more silver nitrate and μTBS decrease than the MDP group (P < 0.05). These results indicated that MPS had a weak interaction with collagen that enhanced its surface negative charge and hydrophobicity without adversely affecting dentin bonding. However, compared to MDP alone, mixing MDP with MPS impaired their effectiveness and made the dentin bonding unstable.
“Go Green” manoeuvre is gaining prominence due to environmental sustainability, where chemical consumption needs to be subjugated. In order to alleviate this issue, industries have adopted the use of multifunctional chemicals as an alternative to mono-functional agents. The global interest in cyclic carbonate is emerging because of its solvent-cum-reagent characteristic. In this work, we have attempted to use “Propylene Carbonate (PC)” as a green multifunctional agent in the different unit processes of leather making. The antimicrobial activity of PC against gram-negative (Bacillus subtilis, Staphylococcus aureus), gram-positive (Escherichia coli, Klebsiella pneumonia) bacterial strains, and mixed culture prepared from putrefied skin extract enables to use of PC in the preservation process as an alternative to common salt (NaCl). The goatskins treated with 15% PC can be stored for up to 2 months without any putrefaction. Further, the developed method reduces the 85% of TDS load from the soaking process and also completely eliminates the generation of contaminated salt. Spectroscopic investigation and molecular dynamic simulation studies clearly indicate that the structural stability of collagen is not affected in the presence of PC. However, it inhibits the activity of collagenase on collagen. The use of PC as an alkali neutralizing agent in the deliming process completely avoids the generation of toxic ammonium gas. The possibility of using PC as an antifungal agent in the chrome tanning process has been attempted. The wet-blue leather treated with 3% PC has been preserved for 30 days without any fungal attack. Hence, the application of PC in various unit processes has a great potential in reducing chemical consumption thereby leading to sustainable leather manufacturing.
Objectives
To investigate the relationship between magnetization transfer (MT) imaging and tissue macromolecules in high grade serous ovarian cancer (HGSOC) and whether MT ratio (MTR) changes following neoadjuvant chemotherapy (NACT).
Methods
This was a prospective observational study. Twelve HGSOC patients were imaged before treatment. MTR was compared to quantified tissue histology and immunohistochemistry. For a subset of patients (n = 5), MT imaging was repeated after NACT. The Shapiro-Wilk test was used to assess for normality of data and Spearman’s rank-order or Pearson’s correlation tests were then used to compare MTR with tissue quantifications. The Wilcoxon signed-rank test was used to assess for changes in MTR after treatment.
Results
Treatment-naïve tumour MTR was 21.9±3.1% (mean ± S.D.). MTR had a positive correlation with cellularity, rho = 0.56 (p < 0.05) and a negative correlation with tumour volume, rho = −0.72 (p = 0.01). MTR did not correlate with the extracellular proteins, collagen IV or laminin (p = 0.40 and p = 0.90). For those patients imaged before and after NACT, an increase in MTR was observed in each case with mean MTR 20.6±3.1% (median 21.1) pre-treatment and 25.6±3.4% (median 26.5) post-treatment (p = 0.06).
Conclusion
In treatment-naïve HGSOC, MTR is associated with cellularity, possibly reflecting intracellular macromolecular concentration. MT may also detect the HGSOC response to NACT, however larger studies are required to validate this finding.
Advances in knowledge
MTR in HGSOC is influenced by cellularity. This may be applied to assess for cell changes following treatment.
Cortical bone tissue, primarily composed of collagen, hydroxyapatite, and water, is a strong and tough natural, structural biomaterial. The integrity of the collagenous phase (native triple helix vs. damaged/denatured coil) has previously been correlated via various means, including hydrothermal isometric tension testing and FTIR and Raman spectroscopy, with the capability of cortical bone to undergo stable fracture. Collagen is a relatively stable protein, requiring around 70 J/g to thermally denature its native triple helix structure, through the melting of hydrogen bonds. It is widely thought that bone collagen molecules denature (unravel) during fracture, acting as a molecular-scale mechanical toughening mechanism, but this has not been empirically demonstrated to date. A new technology, fluorescently-labelled collagen hybridizing peptides (F–CHP), enables imaging that specifically detects denatured collagen. This provides an opportunity to empirically test whether bone collagen molecules do denature during bone fracture. Here, F–CHP was used to stain fracture surfaces produced by transverse Mode-I fracture of chevron-notched bovine and human cortical bone beams. The fracture surfaces demonstrated increased staining, above the level of rigorous paired controls, and the staining directly correlated with the work-to-fracture (WFx) of bovine bone beams. This increased denaturation signal was also constrained to a rough textured region visible on the fracture surface, which is known to correspond with stable tearing. Similar staining was also detected on the fracture surfaces of human cortical bone. Increased staining was not detected on the fracture surfaces of specimens that were dehydrated prior to fracture, suggesting a role for water in the denaturation process. This study provides the first empirical evidence of bone collagen denaturation resulting from cortical bone fracture and extends our understanding of this mechanism towards the mechanical performance of cortical bone.
The 3D bioprinting of cell-incorporated gels is a promising direction in tissue engineering applications. Collagen-based hydrogels, due to their similarity to extracellular matrix tissue, can be a good candidate for bioink and 3D bioprinting applications. However, low hydrogel concentrations of hydrogel (<10 mg/mL) provide insufficient structural support and, in highly concentrated gels, cell proliferation is reduced. In this study, we showed that it is possible to print highly concentrated collagen hydrogels with incorporated cells, where the viability of the cells in the gel remains very good. This can be achieved simply by optimizing the properties of the bioink, particularly the gel composition and pH modification, as well as by optimizing the printing parameters. The bioink composed of porcine collagen hydrogel with a collagen concentration of 20 mg/mL was tested, while the final bioink collagen concentration was 10 mg/mL. This bioink was modified with 0, 5, 9, 13, 17 and 20 μL/mL of 1M NaOH solution, which affected the resulting pH and gelling time. Cylindrical samples based on the given bioink, with the incorporation of porcine adipose-derived stromal cells, were printed with a custom 3D bioprinter. These constructs were cultivated in static conditions for 6 h, and 3 and 5 days. Cell viability and morphology were evaluated. Mechanical properties were evaluated by means of a compression test. Our results showed that optimal composition and the addition of 13 μL NaOH per mL of bioink adjusted the pH of the bioink enough to allow cells to grow and divide. This modification also contributed to a higher elastic modulus, making it possible to print structures up to several millimeters with sufficient mechanical resistance. We optimized the bioprinter parameters for printing low-viscosity bioinks. With this experiment, we showed that a high concentration of collagen gels may not be a limiting factor for cell proliferation.
More than 70 mutations in the two structural genes for type I procollagen (COL1A1 and COL1A2) have been found in probands with osteogenesis imperfecta, a heritable disease of children characterized by fragility of bone and other tissues rich in type I collagen. The mutations include deletions, insertions, RNA splicing mutations, and single-base substitutions that convert a codon for glycine to a codon for an amino acid with a bulkier side chain. With a few exceptions, the most severe phenotypes of the disease are explained largely by synthesis of structurally defective pro alpha chains of type I procollagen that either interfere with the folding of the triple helix or with self-assembly of collagen into fibrils. The results emphasize the extent to which the zipperlike folding of the collagen triple helix and the self-assembly of collagen fibrils depend on the principle of nucleated growth whereby a few subunits form a nucleus and the nucleus is then propagated to generate a large structure with a precisely defined architecture. The principle of nucleated growth is a highly efficient mechanism for the assembly of large structures, but biological systems that depend extensively on nucleated growth are highly vulnerable to mutations that cause synthesis of structurally abnormal but partially functional subunits. Recently, several mutations in three other collagen genes (COL2A1, COL3A1, and COL4A5) have been found in probands with genetic diseases involving tissues rich in these collagens. Most of the probands have rare genetic diseases but a few appear to have phenotypes that are difficult to distinguish from more common disorders such as osteoarthritis, osteoporosis, and aortic aneurysms. Therefore, the results suggest that mutations in procollagen genes may cause a wide spectrum of both rare and common human diseases.
A 2:2 complex of proflavine and deoxycytidylyl-3′,5′-guanosine has been crystallized and its structure determined by x-ray
crystallography.
The two dinucleoside phosphate strands form self complementary duplexes with Watson Crick hydrogen bonds. One proflavin is
asymmetrically intercalated between the base pairs and the other is stacked above them. The conformations of the nucleotides
are unusual in that one strand has C3′,C2′ endo mixed sugar puckering and the other has C3′,C3′ endo deoxyribose sugars. These
results show that the conformation of the 3′ sugar is of secondary importance to the intercalated geometry.
We report direct measurements of force vs. separation between self-assembled proteins. These forces are observed between collagen triple helices in native and reconstituted fibers. They are a combination of a short-range repulsion, which varies exponentially over at least five decay lengths, and an inferred, longer-ranged attraction responsible for spontaneous assembly. From 5 degrees C to 35 degrees C the relative contribution of the attraction to the net force increases with temperature. These forces are strikingly similar to the "hydration" forces measured between several other linear macromolecules (DNA, polysaccharides) and between lipid bilayer membranes. The decay length of the repulsive force agrees well with a theoretical estimate based on axial periodicity of the triple helix, suggesting another connection between molecular architecture and protein-protein interaction.
Macrophage scavenger receptors have been implicated in the development of atherosclerosis and other macrophage-associated functions, including host defense. The mechanism by which these receptors bind a wide array of polyanions, such as acetylated low density lipoprotein (Ac-LDL), with high affinity has not yet been elucidated; however, it has been proposed that the positively charged extracellular collagenous domain of scavenger receptors plays a key role in ligand binding. To test this proposal, we generated truncation mutants of the bovine and murine scavenger receptors and studied their expression in transiently transfected COS cells. These mutants contain only 8 (bovine) or 5 (murine) of the 24 Gly-X-Y tripeptide repeats found in the collagenous domains of the full-length receptors. Immunochemical analyses established that the truncation of the bovine scavenger receptor did not interfere significantly with its synthesis, trimerization, post-translational processing, intracellular transport, surface expression, or stability. However, unlike their full-length counterparts, the truncated bovine and murine receptors were unable to bind Ac-LDL. Thus, the collagenous domain was necessary for normal ligand binding. In addition, cotransfection of the expression vector for the truncated bovine scavenger receptor with that for the full-length receptor resulted in dramatically reduced activity of the full-length construct (dominant negative effect). A ligand bead-binding assay was used to show that the isolated collagenous domain from a different protein, complement component C1q, could bind a wide variety of polyanions with a specificity which was similar, but not identical, to that of scavenger receptors. These results suggest that the collagenous domain of the scavenger receptor is both necessary and sufficient to determine the broad binding specificity that characterizes this unusual receptor. Scavenger receptors and C1q, along with the mannose-binding protein, conglutinin, and lung surfactant apoprotein A, help define a set of proteins which all contain short collagenous domains and which all appear to participate in host defense. Their short collagenous domains may contribute significantly to their host-defense functions.
The water structure has been analyzed for a model of the protein crambin refined against 0.945-A x-ray diffraction data. Crystals contain 32% solvent by volume, and 77% of the solvent molecules have been located-i.e., 2 ethanol molecules and 64 water molecules with 10-14 alternate positions. Many water oxygen atoms found form chains between polar groups on the surface of the protein. However, a cluster of pentagonal arrays made up of 16 water molecules sits at a hydrophobic, intermolecular cleft and forms a cap around the methyl group of leucine-18. Several waters in the cluster are hydrogen-bonded directly to the protein. Additional closed circular arrays, which include both protein atoms and other water oxygen atoms, form next to the central cluster. This water array stretches in the b lattice direction between groups of three ionic side chains.
Hydration is so important for the structural, physical and biological properties of the proteins, that it has been studied by a variety of methods such as Raman, IR and NMR spectroscopy, calorimetry, gravimetry, molecular dynamics [621, 622, 810, 827–834].
The chapter discusses the protein stability with emphasis on compact globular proteins representing a single cooperative system. All the small compact globular proteins represent cooperative systems; they exhibit an extreme cooperativity that integrates the whole of their structure into a single structural unit. The large proteins, to which fibrillar proteins are also related, do not present single cooperative systems, but are subdivided into definite cooperative subsystems—structural blocks or domains. The advances in studying the stability of complicated proteins are connected with two methodical achievements: (1) the appearance of the precise scanning microcalorimetric technique, which affords reliable information on the heat capacity function of proteins in a broad temperature range; and (2) realization of the fact that the complicated heat effect of disruption of a complex macromolecular structure can be analyzed thermodynamically. The thermodynamic specificity of collagen has been considered. The volume of globular proteins does not increase at denaturation but decreases, as seen from their ability to denature under high pressure. The results of calorimetric studies are discussed, presenting the specific melting enthalpy of various protein structures—globular proteins, double-stranded coiled coils, and triplestranded coiled coils. The practical importance of thermodynamic studies of protein stability—that is, its importance not only for understanding the principles of organization of these molecules, but just for obtaining structural information on the domain level is emphasized.
Bond-length and bond-angle parameters are derived from a statistical survey of X-ray structures of small compounds from the Cambridge Structural Database. The side chains of the common amino acids and the polypeptide backbone were represented by appropriate chemical fragments taken from the Database. Average bond lengths and bond angles are determined from the resulting samples and the sample standard deviations provide information regarding the expected variability of the average values which can be parametrized as force constants. These parameters are ideally suited for the refinement of protein structures determined by X-ray crystallography since they are derived from X-ray structures, are accurate to within the deviations from target values suggested for X-ray structure refinement and use force constants which directly reflect the variability or uncertainty of the average values. Tests of refinement of the structures of BPTI and phycocyanin demonstrate the integrity of the parameters and comparisons of equivalent refinements with XPLOR parameters show improvement in R factors and geometry statistics.
In order to analyze the directionality of hydrogen bonding to oxygen atoms the Cambridge Crystallographic Data File was searched for O. . . X (X equals N, O) intermolecular contacts at less than 3 A in structures containing ether, ketone, epoxide, enone, and ester groups. The results are represented as scatterplots. However, for further clarity, they are also represented as diffuse (probability) densities obtained by superposing spherical atomic electron density functions with a parameterizable temperature factor on each point in the scatterplot and contouring the maps at a height proportional to the number of data points. In all systems the largest concentration of hydrogen-bonded X groups lay in the direction commonly ascribed to lone pairs, but generally only one such X group per oxygen atom.
At any fixed temperature between -90 and +30°C the partial molar heat capacity of water absorbed in collagen fibers is independent of concentration from 0 to 0.5 g of water per gram of dry collagen. This result is in disagreement with all two-state models proposed. The high value of 22 cal deg-1 mol-1 for the partial molar heat capacity of water decreases gradually to a value of 9 cal deg-1 mol-1 at -90°C, a value approaching that of ice. We conclude that water becomes vitreous in the vicinity of -100°C. Our results are in agreement with the previously proposed one-state, liquidlike, model for water absorbed in collagen. According to this model water molecules are hydrogen bonded in long chains that diffuse through the interstices in liquidlike fashion.
Nuclear magnetic resonance and dielectric data on hydrated collagen are interpreted in terms of Ramachandran's hydration model. It is found that all data are compatible with this model, indicating two specific binding sites per three amino acids in the threefold collagen helix. Sorption data have been interpreted according to the multilayer theory of Guggenheim and used to derive the fraction of bound water in the primary sites. From magnetic resonance anisotropies structural details of the position of the water molecules can be derived under the assumption that both sites are equally occupied. The residence time of a water molecule in one of these sites in moderately hydrated collagen (45 g H2O/100 g collagen) is 1.2 × 10−6 sec. The remainder of the water is weakly bound and consists of rapidly exchanging species with rotational correlation time shorter than 10−10 sec. The sites are 50% occupied at a water content of 10 g/100 g collagen and may contribute significantly to the stability of the collagen threefold helix.
In this chapter, we discuss the current understanding of the family of matrix macromolecules that are classified as collagens. Reference is made to reviews and recent papers to provide the reader with a way into the relevant literature rather than an exhaustive survey of the bibliography.
The collectins are a group of mammalian lectins containing collagen-like regions. They include mannan binding protein, bovine conglutinin, lung surfactant protein A, lung surfactant protein D, and a newly discovered bovine protein named collectin-43. These proteins share a very similar modular domain composition and overall 3-dimensional structure. They also appear to play similar biological roles in the preimmune defense against microorganisms in both serum and lung surfactant. The close evolutionary relationship between the collectins is further emphasized by a common pattern of exons in their genomic structures and the presence of a gene cluster on chromosome 10 in humans that contains the genes known for the human collectins. Studies on the structure/function relationships within the collectins could provide insight into the properties of a growing number of proteins also containing collagenous regions such as Clq, the hibernation protein, the α- and β-ficolins, as well as the membrane acetylcholinesterase and the macrophage scavenger receptor.
Two different collagens were isolated and characterized from the body walls of the vestimentiferan tube worm Riftia pachyptila and the annelid Alvinella pompejana, both living around hydrothermal vents at a depth of 2600 m. The acid-soluble cuticle collagens consisted of a long triple helix (2·4 μm for Alvinella, 1·5 μm for Riftia) terminating into a globular domain. Molecular masses of 2600 and 1700 kDa, respectively, were estimated from their dimensions. The two cuticle collagens were also quite different in amino acid composition, in agreement with their different supramolecular organizations within tissues. Interstitial collagens corresponding to cross-striated fibrils underneath the epidermal cells could be solubilized by digestion with pepsin and consisted of a single α-chain. They were similar in molecular mass (340 kDa) and length (280 nm) but differed in composition and banding patterns of segment-long-spacing fibrils. This implicates significant sequence differences also in comparison to fibril-forming vertebrate collagens, although all form typical quarter-staggered fibrils. The thermal stability of the worm collagens was, with one exception (interstitial collagen of Riftia), in the range of mammalian and bird collagens (37 to 46°C), and thus distinctly above that of shallow sea water annelids. Yet, their 4-hydroxyproline contents were not directly correlated to this stability. About 20% of Riftia collagen α-chain sequence was elucidated by Edman degradation and showed typical Gly-X-Y repeats but only a limited homology (45 to 58% identity) to fibril-forming vertebrate collagens. A single triplet imperfection and the variable hydroxylation of proline in the X position were additional unique features. It suggests that this collagen represents an ancestral form of fibril-forming collagens not directly corresponding to an individual fibril-forming collagen type of vertebrates.
This paper describes in detail our work on the structure of collagen which we have already outlined elsewhere (Rich & Crick, 1955). The main substance of the paper is:(1)a demonstration that, given certain assumptions, only two basic types of structures are possible for collagen;(2)detailed work on the coordinates and Fourier transforms of one of these models (collagen II), and a comparison between these predictions and the observed X-ray diffraction data.
(Pro-Hyp-Gly)5 and (Pro-Hyp-Gly)10 were synthesized by repeated condensation of tert-butyloxycarbonyl(Boc)-Pro-Hyp[benzyl(Bzl)]-Gly on a Merrifield resin followed by cleavage of the peptides from the resin with HF. The solution properties of the peptides were then compared with those of (Pro-Pro-Gly)5 and (Pro-Pro-Gly)10 synthesized previously. The peptides containing hydroxyproline were similar to those that did not, in that they formed triple-helical structures analogous to the triple-helical structure of collagen. At low temperatures the peptides showed a high degree of negative optical rotation and there was a relatively sharp change in optical rotation as the temperature was increased. The was about 5 °C for (Pro-Hyp-Gly)5 and about 58 °C for (Pro-Hyp-Gly)10. Ultracentrifugation studies were consistent with the conclusion that the temperature-dependent changes in optical rotation reflected a transition between triple-stranded and single-stranded forms of the peptides. The major difference between the peptides containing hydroxyproline and those that did not was that the values for the hydroxyproline-containing peptides were higher by about 35 °C. The results indicated that the hydroxyprolyl residues preceding glycyl residues in the peptides stabilize the triple-helical structure, and they suggested therefore that hydroxyproline may perform a similar function in collagen.
Quantitative X-ray diffraction data have been collected from stretched kangaroo tail tendon and used to test models for the conformation of the polypeptide chains in the collagen molecule. The magnitude of the unit twist of the molecular helix was estimated to be 107.1 ° ± 0.6 °, which is close to the value expected for a helix with ten units in three turns. The intensity data were used to carry out a linked-atom least-squares refinement of models based on two possible interchain hydrogen bonding schemes suggested by Rich & Crick (1955, 1961). No stereochemically acceptable solution could be found for the hydrogen bonding scheme of model I, but a stereochemically satisfactory solution was found for the scheme of model II which gave a crystallographic R factor of 0.272.
The interaction of water with collagenous tissue was investigated using dynamic mechanical spectroscopy and cryogenic X-ray techniques. The loss spectrum was found to be very sensitive to water which is highly associated with the macromolecule. Two water-sensitive loss peaks were observed below 0°C: the β2 or “water dispersion” at 150°K and the β1 at 200°K which is attributed to motion of polar side chains. Changes in peak temperature and intensity were not continuous with water content, but exhibited regimes in behavior which were associated with two types of nonfreezable water, structural and bound water. In cryogenic X-ray experiments, specimens which contained some freezable water exhibited reflections identified with the cubic form of ice. These ice crystals underwent an irreversible transition to the more common hexagonal form when warmed above 200°K. On the basis of these experiments, a model for the hydration of native collagenous tissue was proposed.
The collagen-like peptides (L-Pro-L-Pro-Gly)n and (L-Pro-L-Hyp-Gly)n with n = 5 and 10, were examined in terms of their triple helix ⇌ coil transitions in aqueous and nonaqueous solvents. The peptides were soluble in 1,2-propanediol containing 3% acetic acid and they were found to form triple-helical structures in this solvent system. The water content of the solvent system and the amount of water bound to the peptides were assayed by equilibrating the solvent with molecular sieves and carrying out Karl Fischer titrations on the solvent phase. After the solvent was dehydrated, much less than one molecule of water per tripeptide unit was bound to the peptides. Since the peptides remained in a triple-helical conformation, the results indicated that water was not an essential component of the triple-helical structure. Comparison of peptides with the same chain length demonstrated that the presence of hydroxyproline increased the thermal stability of the triple helix even under anhydrous conditions. The results, therefore, did not support recent hypotheses that hydroxyproline stabilizes the triple helix of collagen and collagen-like peptides by a specific interaction with water molecules. Analysis of the thermal transition curves in several solvent systems showed that although the peptides containing hydroxyproline had tm values which were 18.6° to 32.7°C higher, the effect of hydroxyproline on ΔG was only 0.1 to 0.3 kcal per tripeptide unit at 25°C. The results suggested, therefore, that the influence of hydroxyproline on helical stability may be explained by intrinsic effects such as dipole–dipole interactions or by changes in the solvation of the peptides by alcohol, acetic acid, and water. A direct calorimetric measurement of the transition enthalpy for (L-Pro-L-Pro-Gly)n in 3% or 10% acetic acid gave a value of −1.84 kcal per tripeptide unit for the coil-to-helix transition. From the value for enthalpy and from data on the effects of different chain lengths on the thermal transition, it was calculated that the apparent free energy for nucleation was +5 kcal/mol at 25°C (apparent nucleation parameter = 2 × 10−4M−2). The value was dependent on solvent and on chemical modification of end groups.
A solution of the problem of topology of a hydrogen bond net in a triple helix of collagen is suggested on the basis of an analysis of thermodynamic data on denaturation of phylogenetically different collagen [T. V. Burjanadze (1982), Vol. 21, pp. 1489-1501; T. V. Burjanadze, E. I. Tiktopulo, and P. L. Privalov (1987), Dokl. Akad. Nauk. USSR, Vol. 293, pp. 720-724] as well as on the earlier evaluation of the energy of the OH group of the 4-hydroxyproline bond [A. R. Ward and P. Mason (1973), Journal of Molecular Biology, Vol. 29, pp. 431-435]. It is shown that only the water-bridged collagen structure [G. N. Ramachandran and R. Chandrasekharan (1968), Biopolymers, Vol. 6, pp. 1649-1661; G. N. Ramachandran, M. Bansal, and R. S. Bhatnagar (1973), Biochimica Biophysica Acta, Vol. 322, pp. 166-171; M. Bansal, C. Ramakrishnan, and G. N. Ramachandran (1975), Proceedings of the Indian Academy of Sciences, Vol. 82, pp. 152-164] can explain both the change of thermostability upon proline hydroxylation [J. Rosenbloom, M. Harsch, and S. Jimenez (1973), Archives of Biochemistry and Biophysics, Vol. 158, pp. 478-484] and its phylogenetic change [T. V. Burjanadze (1982)].
Nuclear magnetic relaxation times were measured in collagen tissue when varying the orientation of the fiber with respect to the static field. T1 was found to be only slightly dependent on theta, the fiber-to-field angle, but T2 was very sensitive to the orientation, with a maximum value at the magic angle. The transverse decay curves were multiexponential. Their deconvolution displayed four components; the ones that decayed most slowly were almost independent of theta, but the two fastest ones showed a strong angular dependence that was interpreted with a cross-relaxation model. Quadrupolar dips were visible in the 1/T1 dispersion curves. These dips were independent of theta, so that the magnetization transfer could also be assumed to be independent of the fiber orientation. Finally, each component was assigned to a fraction of protons localized in the macromolecular structure and characterized by particular dynamics. The model of Woessner was applied to the water molecules tightly bound into the macromolecules, which resulted in a dynamical description of this water fraction. This description is compatible with the two-sites model of Ramachandran based on x-ray diffraction and with the extensive studies of Berendsen. However, the important indications obtained from the deconvolution lead to a less static representation of the tissue.
The macrophage scavenger receptor is a trimeric membrane glycoprotein with unusual ligand-binding properties which has been implicated in the development of atherosclerosis. The trimeric structure of the bovine type I scavenger receptor, deduced by complementary DNA cloning, contains three extracellular C-terminal cysteine-rich domains connected to the transmembrane domain by a long fibrous stalk. This stalk structure, composed of an alpha-helical coiled coil and a collagen-like triple helix, has not previously been observed in an integral membrane protein.
Crystals of [Phe4 Val6] antamanide (cyclic [ValProProPhePhe]2) grown from dioxane/H2O, with space group P21212 and cell parameters a = 15.099(4), b = 22.008(5) and c = 11.024(3) A, are almost identical to crystals grown from H2O/acetone, the structure of which was determined a number of years ago. Per peptide molecule there are the equivalent of 12 water molecules occupying 16 sites in both crystals; however, in the new investigation a number of water molecules present at one-half occupancy have been found in different positions than in the earlier analysis. The interpretation of the hydrogen bonding between peptide/water and between water/water is much more satisfactory. Pentagonal water assemblies are present in the solvent channel. There is a distinct indication of the occurrence of a bifurcated bond between two water molecules, as well as the presence of three-center hydrogen bonds joining three water molecules. This may be the first experimental example of a bifurcated bond between two water molecules.
The amino acid compositions of Metridium-, Lumbricus- and Mytilus-collagens corresponded to those of the lowest vertebrates: the content of imino acids was low and that of serine and threonine high. Metridium-collagen also contained cystine and large amounts of hydroxylysine. In Lumbricus-collagen the hydroxylation of proline was almost complete and the total content of hydroxy amino acids thus exceptionally high.
The solubility of collagens from the invertebrates varied in wide range. In some cases, e. g. the byssus apparatus of Mytilus, the insolubility seems to depend on the stabilizing effect of the adjacent materials which is manifested also in the unique thermal shrinking temperature of 90°.
The starch-gel electrophoretic patterns were studied on soluble collagens from the following species: octopus, Loligo, Lumbricus, Diphyllobothrium, Mytilus, and Metridium. A single α-component pattern, resembling that from lamprey, was observed in all the cases.
A series of nine procollagen samples in which the hydroxyproline content varied from <1% to 44% of the total imino acids was prepared by incubating embryonic chick tendon cells with varying concentrations of α,α′-dipyridyl, an inhibitor of proline hydroxylase. The thermal stability of these procollagen preparations was then investigated by using pepsin digestion at different temperatures as an enzymatic probe of conformation. Using this technique, the denaturation temperature of the procollagen was found to be directly proportional to the hydroxyproline content. A denaturation temperature of 23.5 °C was found for the unhydroxylated procollagen and 37.9 °C for fully hydroxylated procollagen. These results suggest that hydroxyproline is crucial to the thermal stability of the collagen triple helix. They also imply that unhydroxylated molecules are not triple helical within the cell at 37 °C and that triple helix formation may be necessary for normal secretion.
The possibility of hydroxyproline residues stabilizing the collagen triple-helical structure by the formation of additional hydrogen bonds through their γ-hydroxyl group has been studied from structural considerations. It is not possible for this hydroxyl group to form a direct hydrogen bond with a suitable group in a neighbouring chain of the triple-helical protofibril. However, in the modified one-bonded structure, which is stabilized by additional hydrogen bonds being formed through water molecules as intermediaries (put forward in 1968 by Ramachandran, G. N. and Chandrasekharan, R.), it is found that the γ-hydroxyl group of hydroxyproline can form a good hydrogen bond with the water oxygen as acceptor, the hydrogen bond length being 2.82 Å. It is proposed that, in addition to stabilizing the collagen triple-helical structure due to the stereochemical properties of the pyrrolidine ring, hydroxyproline gives added stability by the formation of an extra hydrogen bond. Experimental studies on the determination of shrinkage and denaturation temperatures of native collagen and its synthetic analogues, as a function of their hydroxyproline content, are being undertaken to test this hypothesis.
If the collagen triple helix is so built as to have one set of NH ⃛ O hydrogen bonds of the type N3H3(A) ⃛ O2(B), then it is possible to have a linkage between N1H1(B) and O1(A) through the intermediary of a water molecule with an oxygen O leading to the formation of the hydrogen bonds N1(B) ⃛ O and O (A). In the same configuration, another water molecule with an oxygen O can link two earbonyl oxygens of chains A and B forming the hydrogen bonds O O1(A) and O O0 (B). The two water oxygens also become receptors at the same time for CH ⃛ O hydrogen bonds. Thus, the neighboring chains in the triple helix are held together by secondary valence bond linkages occurring regularly sit intervals of about 3 Å along the length of the protofibril. The additional water molecules occur on the periphery of the proto-fibril and will contribute their full share towards stabilizing the structure in the solid state. In solution, they will be disturbed by the medium unless they are protected by long side groups. It appears that this type of two-bonded structure, in which one NH ⃛ O bond is to a water molecule, can explain several observations on the stability and hydrogen exchange properties of collagen itself and related synthetic polypeptides. The nature of the water bonds and their strength are found to be better in the one-bonded structure proposed from Madras than in the one having the coordinates of Rich and Crick.
COLLAGEN is a ``young'' protein present only in multicellular organisms, and the number of acceptable mutations of the amino-acid composition is restricted by the structural requirements of the secondary structure. In what follows we summarize some of our work1-5 during the past 6 years in which we have compared collagen from different species.
Brillouin scattering was used to determine the longitudinal acoustic velocity along the axis of collagen fibers as the relative humidity decreased from 95 to 0%. Between 0 and 86%, the velocity decreased linearly at a modest rate. Above 86%, the decrease was steep. The changing mass of the absorbed water accounts for most of the velocity change in the lower humidity range. The remaining deviation is probably due to variation of the elastic coupling between molecules. At low humidity, the adsorbed water is known to be attached to the collagen molecules, whereas the additionally adsorbed water at high humidity has the properties of bulk liquid. The high-humidity sonic velocity variation is ascribed to the presence of free water. It is possible to identify the five water regimes of Pineri et al. [(1978) Biopolymers17, 2799–2815] with the variations of the sonic velocity with water content.
(Pro-Pro-Gly)10 forms single crystals, providing X-ray diffraction data to 0.22 nm resolution. In the crystals, the polypeptides form triplexes that aggregate end-to-end in quasi-infinite helices with axial translation per tripeptide h = 0.287 nm and the corresponding rotation t = −102.9 °. The structure, which may be an allomorph of collagen, has been refined by the linked-atom least-squares procedure. In addition, three water molecules per tripeptide have been detected by Fourier difference syntheses. One of them forms an intrachain hydrogen-bonded bridge O(Pro2) - - - W - - - O(Gly). There are also interchain hydrogen bonds (Gly)NH - - - O(Pro1) within the triplex.
X-ray diffraction analysis of crystals of the intercalative complex between the deoxyribonucleoside phosphate d(CpG) and the mutagen proflavine shows a highly structured arrangement of water molecules linked together by newtworks of hydrogen bonds to form four edge-linked pentagons per asymmetric unit. These pentagons have a general role in maximizing hydrogen bonding at 3.4-A intervals. The conformation of the deoxyribose sugar ring at the 3' end of one strand can depend on its local aqueous environment.
The structure of a protein triple helix has been determined at 1.9 angstrom resolution by x-ray crystallographic studies of a collagen-like peptide containing a single substitution of the consensus sequence. This peptide adopts a triple-helical structure that confirms the basic features determined from fiber diffraction studies on collagen: supercoiling of polyproline II helices and interchain hydrogen bonding that follows the model II of Rich and Crick. In addition, the structure provides new information concerning the nature of this protein fold. Each triple helix is surrounded by a cylinder of hydration, with an extensive hydrogen bonding network between water molecules and peptide acceptor groups. Hydroxyproline residues have a critical role in this water network. The interaxial spacing of triple helices in the crystal is similar to that in collagen fibrils, and the water networks linking adjacent triple helices in the crystal structure are likely to be present in connective tissues. The breaking of the repeating (X-Y-Gly)n pattern by a Gly-->Ala substitution results in a subtle alteration of the conformation, with a local untwisting of the triple helix. At the substitution site, direct interchain hydrogen bonds are replaced with interstitial water bridges between the peptide groups. Similar conformational changes may occur in Gly-->X mutated collagens responsible for the diseases osteogenesis imperfecta, chondrodysplasias, and Ehlers-Danlos syndrome IV.
A novel murine plasma membrane protein has been identified in subpopulations of macrophages. It has an intracellular N-terminal domain, a transmembrane domain, and an extracellular region with a short spacer, an 89 Gly-Xaa-Yaa repeat-containing collagenous domain, and a C-terminal cysteine-rich domain. In situ hybridization and immunohistochemical staining have localized the protein to a subset of macrophages in the marginal zone of the spleen and the medullary cord of lymph nodes. No expression was observed in macrophages of liver or lung. Transfected COS cells synthesized a native trimeric plasma membrane protein that bound labeled bacteria and acetylated LDL, but not yeast or Ficoll. The results suggest that the novel protein is a macrophage-specific membrane receptor with a role in host defense, as it shows postnatal expression in macrophages, which are considered responsible for the binding of bacterial antigens and phagocytosis.
A new method to analyze the distribution of water molecules around the bases in DNA is presented. This method relies on the notion of a "hydrated building block," which represents the joint observed hydration around all bases of a particular type, in structures of a particular conformation type. The hydrated building blocks were constructed using atomic coordinates from 40 structures contained in the Nucleic Acid Database. Pseudoelectron densities were calculated for water molecules in each hydrated building block using standard crystallographic procedures. The electron densities were fitted to obtain "average building blocks," which represent bases with waters only at average or probable positions. Both types of building blocks were used to construct models of hydrated DNA oligomers. The essential features of the solvent structure around d(CGCGAATTCGCG)2 in the B form and d(CGCGCG)2 in the Z form were reproduced.
X-PLOR Manual, Version 3.1
- A T Brunger
Brunger, A.T. (1992). X-PLOR Manual, Version 3.1. Yale University
Press, New Haven, CT.
- I L Karle
Karle, I.L. (1980). dioxane. nt. J. Pept. Protein Res. 28, 6-12.