Marco Moracci

University of Naples Federico II | UNINA · Department of Biology

On August 26, 2023, we bid farewell to Professor Mosè Rossi, a distinguished figure in the field of enzymology and a beloved member of the International Society for Extremophiles since 1993. Born in Castellabate (Salerno) in 1938, Professor Rossi embarked on a remarkable journey in the world of science.

The space race is entering a new era of exploration, in which the number of robotic and human missions to various places in our solar system is rapidly increasing. Despite the recent advances in propulsion and life support technologies, there is a growing need to perform analytical measurements and laboratory experiments across diverse domains of s...

Glycoside hydrolases (GHs) have been employed for industrial and biotechnological purposes and often play an important role in new applications. The red blood cell (RBC) antigen system depends on the composition of oligosaccharides on the surface of erythrocytes, thus defining the ABO blood type classification. Incorrect blood transfusions may lead...

(Hyper)thermophilic archaeal glycosidases are enzymes that catalyze the hydrolysis of glycosidic bonds to break down complex sugars and polysaccharides at high temperatures. These enzymes have an unique structure that allows them to remain stable and functional in extreme environments such as hot springs and hydrothermal vents. This review provides...

Archaea represents the third domain of life, displaying a closer relationship with eukaryotes than bacteria. These microorganisms are valuable model systems for molecular biology and biotechnology. In fact, nowadays, methanogens, halophiles, thermophilic euryarchaeota, and crenarchaeota are the four groups of archaea for which genetic systems have...

The increasing interest for environmentally friendly technologies is driving the transition from fossil-based economy to bioeconomy. A key enabler for circular bioeconomy is to valorize renewable biomasses as feedstock to extract high value-added chemicals. Within this transition the discovery and the use of robust biocatalysts to replace toxic che...

In discovering novel laccase enzymes with interesting catalytic and stability properties, there is still scope for improvement in exploring new sites in Algeria. The bio-prospecting approach allowed the isolation of a novel strain, Streptomyces sp. HBD30 from the Agrioun River, northern Algeria. Following a functional approach, two laccases (LacI a...

Pompe disease is a metabolic myopathy due to acid alpha-glucosidase deficiency. In addition to glycogen storage, secondary dysregulation of cellular functions, such as autophagy and oxidative stress, contributes to the disease pathophysiology. We have tested whether oxidative stress impacts on enzyme replacement therapy with recombinant human alpha...

Genetic code decoding, initially considered to be universal and immutable, is now known to be flexible. In fact, in specific genes, ribosomes deviate from the standard translational rules in a programmed way, a phenomenon globally termed recoding. Translational recoding, which has been found in all domains of life, includes a group of events occurr...

Genetic decoding is flexible, due to programmed deviation of the ribosomes from standard translational rules, globally termed “recoding”. In Archaea, recoding has been unequivocally determined only for termination codon readthrough events that regulate the incorporation of the unusual amino acids selenocysteine and pyrrolysine, and for −1 programme...

In the field of biocatalysis and the development of a bio-based economy, hemicellulases have attracted great interest for various applications in industrial processes. However, the study of the catalytic activity of the lignocellulose-degrading enzymes needs to be improved to achieve the efficient hydrolysis of plant biomasses. In this framework, h...

Background The spore-forming lactic acid bacterium Bacillus coagulans MA-13 has been isolated from canned beans manufacturing and successfully employed for the sustainable production of lactic acid from lignocellulosic biomass. Among lactic acid bacteria, B. coagulans strains are generally recognized as safe (GRAS) for human consumption. Low-cost m...

Background The spore-forming lactic acid bacterium Bacillus coagulans MA-13 has been isolated from canned beans manufacturing and successfully employed for the sustainable production of lactic acid from lignocellulosic biomass. Among lactic acid bacteria, B. coagulans strains are generally recognized as safe (GRAS) for human consumption. Low-cost m...

Pompe disease is an inherited metabolic disorder due to the deficiency of the lysosomal acid α-glucosidase (GAA). The only approved treatment is enzyme replacement therapy with the recombinant enzyme (rhGAA). Further approaches like pharmacological chaperone therapy, based on the stabilising effect induced by small molecules on the target enzyme, c...

Terrestrial hot springs are of great interest to the general public and to scientists alike due to their unique and extreme conditions. These have been sought out by geochemists, astrobiologists, and microbiologists around the globe who are interested in their chemical properties, which provide a strong selective pressure on local microorganisms. D...

The enzymes from hyperthermophilic microorganisms populating volcanic sites represent interesting cases of protein adaptation and biotransformations under conditions where conventional enzymes quickly denature. The difficulties in cultivating extremophiles severely limit access to this class of biocatalysts. To circumvent this problem, we embarked...

Astrochemistry and astrobiology, the fascinating disciplines that strive to unravel the origin of life, have opened unprecedented and unpredicted vistas into exotic compounds as well as extreme or complex reaction conditions of potential relevance for a broad variety of applications. Representative, and so far little explored sources of inspiration...

In the framework of the discovery of new thermophilic enzymes of potential biotechnological interest, we embarked in the characterization of a new thermophilic esterase from the thermophilic bacterium Geobacillus thermodenitrificans. The phy-logenetic analysis of the GTNG_0744 esterase indicated that the sequence belongs to the enterochelin/enterob...

Bacillus coagulans MA-13 is an efficient lactic acid producer which withstands high concentrations of the growth inhibitors formed during the pretreatment of lignocellulosic feedstock. This draft genome sequence is expected to pave the way toward the understanding of mechanisms responsible for the robustness of MA-13 during simultaneous saccharific...

Factors that give enzymes stability, activity, and substrate recognition result from the combination of few weak molecular interactions, which can be difficult to study through rational protein engineering approaches. We used irrational random mutagenesis and in vivo selection to test if a β-glycosidase from the thermoacidophile Saccharolobus solfa...

The reaction mechanism of glycoside hydrolases belonging to family 1 (GH1) of carbohydrate-active enzymes classification, hydrolysing β-O-glycosidic bonds, is well characterised. This family includes several thousands of enzymes with more than 20 different EC numbers depending on the sugar glycone recognised as substrate. Most GH1 β-glycosidases bi...

In this study, a preliminary screening of cellular extracts of S. solfataricus allowed the identification of an α- N -acetylglucosaminidase activity. However, the characterization of the corresponding recombinant enzyme revealed a novel GlcNAc de- N -acetylase, which, in cooperation with the α-glucosidase, catalyzed the hydrolysis of O-α-GlcNAc gly...

It has been reported that the lysosomal enzyme alfa-L-fucosidase-1(FUCA-1) is downregulated in highly aggressive and metastatic types of human tumors.We have analyzed the expression of FUCA-1 in papillary(PTC), anaplastic (ATC) thyroid cancers and in normal thyroid tissues by real time PCR and found that FUCA-1 mRNA was decreased in ATC compared to...

α-L-Rhamnosidases (α-RHAs, EC 3.2.1.40) are glycosyl hydrolases (GHs) hydrolyzing terminal α-L-rhamnose residues from different substrates such as heteropolysaccharides, glycosylated proteins and natural flavonoids. Although the possibility to hydrolyze rhamnose from natural flavonoids has boosted the use of these enzymes in several biotechnologica...

CAZypedia was initiated in 2007 to create a comprehensive, living encyclopedia of the carbohydrate active enzymes (CAZymes) and associated carbohydrate-binding modules involved in the synthesis, modification and degradation of complex carbohydrates. CAZypedia is closely connected with the actively curated CAZy database, which provides a sequence-ba...

Background The Bacillus subtilis spore has long been used to display antigens and enzymes. Spore display can be accomplished by a recombinant and a non-recombinant approach, with the latter proved more efficient than the recombinant one. We used the non-recombinant approach to independently adsorb two thermophilic enzymes, GH10-XA, an endo-1,4-β-xy...

The highly stereocontrolled de novo synthesis of L-NBDNJ (the unnatural enantiomer of the iminosugar drug Miglustat) and a preliminary evaluation of its chaperoning potential are herein reported. L-NBDNJ is able to enhance lysosomal α-glucosidase levels in Pompe disease fibroblasts, either when administered singularly or when co-incubated with the...

Pompe disease, a rare lysosomal storage disease caused by deficiency of the lysosomal acid α-glucosidase (GAA), is characterized by glycogen accumulation, triggering severe secondary cellular damage and resulting in progressive motor handicap and premature death. Numerous disease-causing mutations in the gaa gene have been reported, but the structu...

Geothermally heated regions of Earth, such as terrestrial volcanic areas (fumaroles, hot springs, and geysers) and deep-sea hydrothermal vents, represent a variety of different environments populated by extremophilic archaeal and bacterial microorganisms. Since most of these microbes thriving in such harsh biotopes, they are often recalcitrant to c...

To obtain new insights into community compositions of hyperthermophilic microorganisms, defined as having optimal growth temperatures of 80 °C and above, sediment and water samples were taken from two shallow marine hydrothermal vents (I and II) with temperatures of 100 °C at Vulcano Island, Italy. A combinatorial approach of denaturant gradient ge...

Understanding the formation of biogenic molecules in abiotic conditions is a prerequisite in the origin-of-life studies. Determining the conditions allowing an efficient one-pot synthesis of the largest possible panel of biogenic compounds may shed light on the plausible scenario in which the processes that kick-started life might have occurred. We...

The field of thermophilic microbiology was born in the late 1970s with the pioneering work of Brock (Thermophiles biodiversity, ecology, and evolution. Springer, Boston, pp. 1–9, 2001) and dramatically expanded through the ’80s with the isolation of hyperthermophiles by Stetter (FEMS Microbiol Rev 18:149–158, 1996). The development of SSU rRNA phyl...

Glycans containing α-L-fucose participate in diverse interactions between cells and extracellular matrix. High glycan expression on cell surface is often associated with neoplastic progression. The lysosomal exoenzyme, α-L-fucosidase-1 (FUCA-1) removes fucose residues from glycans. The FUCA-1 gene is down-regulated in highly aggressive and metastat...

We report here on the preparation of a novel α‐thioglycoligase that can be used for the fast and efficient synthesis of α‐ N ‐acetylglucosamine‐based glycosides. Using the α‐ N ‐acetyl‐glucosaminidase from Clostridium perfringens of family GH89 (according to the Carbohydrate Active Enzymes classification) as starting point, we prepared mutants in t...

Chemo-enzymatic synthesis of oligosaccharides exploits the diversity of glycosidases and their ability to promote transglycosylation reactions in parallel with hydrolysis. Methods to increase the transglycosylation/hydrolysis ratio include site-directed mutagenesis and medium modification. The former approach was successful in several cases and has...

In bioeconomy, the development of next generation biorefineries, utilizing energy crops and waste materials, are considered one of the most promising actions toward the sustainable production of fuel and the building blocks for chemicals and polymers with a global market in 2016 of $466.6 billion and a compounded average growth rate of 8.9% between...

Dystroglycan is a highly glycosylated extracellular matrix receptor with essential functions in skeletal muscle and the nervous system. Reduced matrix binding by α-dystroglycan (α-DG) due to perturbed glycosylation is a pathological feature of several forms of muscular dystrophy. Like-acetylglucosaminyltransferase (LARGE) synthesizes the matrix-bin...

Background Biofuels production from plant biomasses is a complex multi-step process with important economic burdens. Several biotechnological approaches have been pursued to reduce biofuels production costs. The aim of the present study was to explore the production in tobacco plastome of three genes encoding (hemi)cellulolytic enzymes from thermop...

The biotransformation of lignocellulose biomasses into fermentable sugars is a very complex procedure including, as one of the most critical steps, the (hemi) cellulose hydrolysis by specific enzymatic cocktails. We explored here, the potential of stable glycoside hydrolases from thermophilic organisms, so far not used in commercial enzymatic prepa...

Glycosynthases are becoming important enzymatic tools for the synthesis of oligosaccharides. Herein, we explore for the first time the synthesis of α-glycoconjugates using a α-glycosynthase in green solvents. Using this biocatalyst, β-Gal-N3 as donor, pNP-Glc and pNP-Man as acceptors, and green co-solvents we obtained high yields and excellent sele...

Hot springs are natural habitats for thermophilic Archaea and Bacteria. In this paper, we present the metagenomic analysis of eight globally distributed terrestrial hot springs from China, Iceland, Italy, Russia, and the USA with a temperature range between 61 and 92 ∘C and pH between 1.8 and 7. A comparison of the biodiversity and community compos...

Here we suggest that the origin of the genetic code, that is to say, the birth of first mRNAs has been triggered by means of a widespread modification of all RNAs (proto-mRNAs and proto-tRNAs), as today observed in the RNA editing and in post-transcriptional modifications of RNAs, which are considered as fossils of this evolutionary stage of the ge...

Pharmacological chaperone therapy is an emerging approach to treat lysosomal storage diseases. Small-molecule chaperones interact with mutant enzymes, favor their correct conformation and enhance their stability. This approach shows significant advantages when compared with existing therapies, particularly in terms of the bioavailability of drugs,...

β-N-acetylhexosaminidases, which are involved in a variety of biological processes including energy metabolism, cell proliferation, signal transduction and in pathogen-related inflammation and autoimmune diseases, are widely distributed in Bacteria and Eukaryotes, but only few examples have been found in Archaea so far. However, N-acetylgluco- and...

The second generation bioethanol represents a main challenge in global efforts to utilize renewable resources rather than fossil fuels. However, the close association of cellulose and hemicelluloses to lignin in the plant cell wall makes it difficult to degrade lignocellulose into fermentable sugars. Consequently, pretreatments are necessary to mak...

Glycosynthases are engineered glycoside hydrolases that in suitable reaction conditions promote the synthesis of oligosaccharides with exquisite stereoselectivity and enhanced regioselectivity, if compared to traditional chemical methods. This approach was demonstrated to be successful in a number of cases including β-glycosynthases acting at the t...

Translational recoding includes a group of events occurring during gene translation, namely stop codon readthrough, programmed ±1 frameshifting, and ribosome bypassing, which have been found in organisms from all domains of life. They serve to regulate protein expression at translational level and represent a relatively less known exception to the...

Pompe disease (PD) is a metabolic myopathy due to the deficiency of the lysosomal enzyme α-glucosidase (GAA). The only approved treatment for this disorder, enzyme replacement with recombinant human GAA (rhGAA), has shown limited therapeutic efficacy in some PD patients. Pharmacological chaperone therapy (PCT), either alone or in combination with e...

The Bacillus subtilis spore has long been used as a surface display system with potential applications in a variety of fields ranging from mucosal vaccine delivery, bioremediation and biocatalyst development. More recently, a non-recombinant approach of spore display has been proposed and heterologous proteins adsorbed on the spore surface. We used...

An interesting approach for the chemo-enzymatic synthesis of carbohydrates is the use of glycosynthases, a class of mutant glycosidases derived from $beta$-glycoside hydrolases obtained by replacement of the active site nucleophile with a non-nucleophilic residue. However, the scarcity of $alpha$-glycosynthases has so far hampered access to the syn...

Oligo-, polysaccharides, and glycoconjugates are a relevant part of the bioactive components of the natural products exploited in therapeutics, diagnostics, food additives, and biomaterials. Glycans are directly involved in important biological processes, such as immunostimulation, anti-inflammatory, antioxidant, and chemoprotectant actions and/or...

Glycosynthases, engineered glycoside hy-drolases that are able to synthesize glycans in quanti-tative yields without hydrolyzing them, are among the most interesting tools for the chemoenzymatic synthesis of carbohydrates that have been made available so far. From their invention in 1998, these enzymes have been developed enormously and dem-onstrat...

Glycosynthases, engineered glycoside hy-drolases that are able to synthesize glycans in quanti-tative yields without hydrolyzing them, are among the most interesting tools for the chemoenzymatic synthesis of carbohydrates that have been made available so far. From their invention in 1998, these enzymes have been developed enormously and dem-onstrat...

Glycosynthases, engineered glycoside hy-drolases that are able to synthesize glycans in quanti-tative yields without hydrolyzing them, are among the most interesting tools for the chemoenzymatic synthesis of carbohydrates that have been made available so far. From their invention in 1998, these enzymes have been developed enormously and dem-onstrat...

Background Carbohydrate-active enzymes (cazymes) are enzymatic activities involved in the hydrolysis, synthesis, recognition, and binding of carbohydrates, thereby contributing to the metabolism and mobilization of sugars and glycoconjugates (glycoproteins and glycolipids). Cazymes play a central role in glycobiology, a relatively new discipline, w...

Hyperthermophilic microorganisms thrives at temperatures higher than 80°C and proteins and enzymes extracted from these sources are optimally stable and active in the presence of temperatures close to the boiling point of water and of other denaturants, i.e. chaotropic agents, pH, organic solvents, detergents, etc. Therefore, hyperstable enzymes ar...

α-Mannosidases, important enzymes in the N-glycan processing and degradation in Eukaryotes, are frequently found in the genome of Bacteria and Archaea in which their function is still largely unknown. The α-mannosidase from the hyperthermophilic Crenarchaeon Sulfolobus solfataricus has been identified and purified from cellular extracts and its gen...

The large-scale production of oligosaccharides is a daunting task, hampering the study of the role of glycans in vivo and the testing of the efficacy of novel glycan-based drugs. Glycosynthases, mutated glycosidases that synthesize oligosaccharides in high yields, are becoming important chemo-enzymatic tools for the production of oligosaccharides....

Glycosidases, the enzymes responsible in nature for the catabolism of carbohydrates, are well-studied catalysts widely used in industrial biotransformations and oligosaccharide synthesis, which are also attractive targets for drug development. Glycosidases from hyperthermophilic organisms (thriving at temperatures > 85 °C) are also interesting mode...

Abstract Carbohydrates serve as structural components and energy sources of cells. More interestingly, however, these biomolecules are involved in a variety of molecular recognition processes in intercellular communication and signal transduction such as cell adhesion, differentiation, development and regulation. For these reasons, great interest h...

Carbohydrate active enzymes (CAZymes) are a large class of enzymes, which build and breakdown the complex carbohydrates of the cell. On the basis of their amino acid sequences they are classified in families and clans that show conserved catalytic mechanism, structure, and active site residues, but may vary in substrate specificity. We report here...

Sequenced genomes often reveal interrupted coding sequences that complicate the annotation process and the subsequent functional characterization of the genes. In the past, interrupted genes were generally considered to be the result of sequencing errors or pseudogenes, that is, gene remnants with little or no biological importance. However, recent...

Fucose-containing oligosaccharides play a central role in physio-pathological events, and fucosylated oligosaccharides have interesting potential applications in biomedicine. No methods for the large-scale production of oligosaccharides are currently available, but the chemo-enzymatic approach is very promising. Glycosynthases, mutated glycosidases...

Glycosynthases are mutant glycosidases, which in the presence of activated glycosides and suitable reaction conditions, synthesize oligosaccharides without hydrolysing them. This feature makes these catalysts promising tools for the large scale synthesis of carbohydrates. However, despite the popularity of the glycosynthetic approach, the number of...

The synthesis of 2-deoxyglycosides and, for the first time, of 2-deoxygalactosides is reported using a thermophilic and thermostable β-glycosyl hydrolase from the archeon Sulfolobus solfataricus and glucal or galactal as donors. The yields observed with alkyl acceptors confirmed that the robustness of the biocatalyst is of great help in designing p...

The β-glycosidase from the hyperthermophilic Archaeon Pyrococcus horikoshii (Phoβ-gly) is a monomeric enzyme with wide substrate specificity belonging to family 1 of glycoside hydrolases classification. Inspection of the three-dimensional structure of the enzyme, recently resolved, showed that Phoβ-gly is membrane bound and that the residues putati...

The Sulfolobus solfataricus β-glycosidase (Sβgly) is a thermostable and thermophilic glycosyl-hydrolase with broad substrate specificity. The enzyme hydrolizes β-D-gluco-, fuco-, and galactosides, and a large number of /Winked glycoside dimers and oligomers, linked β1–3, β1–4, and β1–6, It is able to hydrolize oligosaccharides with up to 5 glucose...

Sulfolobus solfataricus strain Oalpha was previously isolated for its ability to grow on minimal medium supplemented with xylan as a carbon source. The strain exhibited thermostable xylanase activity but several attempts to identify the gene encoding for the activity failed. Further studies showed that the xylanase displayed activity on carboxymeth...

In cell free extracts of the thermoacidophilic gram-positive bacterium Alicyclobacillus acidocaldarius ATCC27009, we have identified beta-gluco- and galactosidase activities showing a specific activity of 0.1 and 12 U/mg, respectively. The two enzymatic activities are associated with different polypeptides and we show here the functional cloning, t...

The thermoacidophilic bacterium Alicyclobacillus acidocaldarius is a rich source of glycoside hydrolases enabling its growth on several di- and polysaccharides. We report here the purification and the characterization of a beta-galactosidase from this source, the cloning of its gene, and the expression and the characterization of the recombinant en...

Glycoside hydrolases form hyperthermophilic archaea are interesting model systems for the study of catalysis at high temperatures and, at the moment, their detailed enzymological characterization is the only approach to define their role in vivo. Family 29 of glycoside hydrolases classification groups alpha-L-fucosidases involved in a variety of bi...

The identification of the determinants of protein thermal stabilization is often pursued by comparing enzymes from hyperthermophiles with their mesophilic counterparts while direct structural comparisons among proteins and enzymes from hyperthermophiles are rather uncommon. Here, oligomeric beta-glycosidases from the hyperthermophilic archaea Sulfo...

The sequences of a number of archaeal genomes have recently been completed, and many more are expected shortly. Consequently, the research of Archaea in general and hyperthermophiles in particular has entered a new phase, with many exciting discoveries to be expected. The wealth of sequence information has already led, and will continue to lead to...

Extremophilic Archaea populate biotopes previously considered inaccessible for life. This feature, and the possibility that they are the extant forms of life closest to the last common ancestor, make these organisms excellent candidates for the study of evolution on Earth and stimulate the exobiological research in planets previously considered tot...

Inhibition of glycosidases has great potential in the quest for highly potent and specific drugs to treat diseases such as diabetes, cancer, and viral infections. One of the most effective ways of designing such compounds is by mimicking the transition state. Here we describe the structural, kinetic, and thermodynamic dissection of binding of two g...

In cell free extracts of the thermoacidophilic gram-positive bacterium Alicyclobacillus acidocaldarius ATCC27009, we have identified beta-gluco- and galactosidase activities showing a specific activity of 0.1 and 12 U/mg, respectively. The two enzymatic activities are associated with different polypeptides and we show here the functional cloning, t...

We have previously shown that the major ion-pairs network of the tetrameric beta-glycosidase from the hyperthermophilic archeon Sulfolobus solfataricus involves more than 16 ion-pairs and hydrogen bonds between several residues from the four subunits and protects the protein from thermal unfolding by sewing the carboxy-termini of the enzyme. We sho...

The standard rules of genetic translational decoding are altered in specific genes by different events that are globally termed recoding. In Archaea recoding has been unequivocally determined so far only for termination codon readthrough events. We study here the mechanism of expression of a gene encoding for a α-l-fucosidase from the archaeon Sulf...

Transglycosylation reactions (autocondensation of the substrate or transfer of the glycon donor moiety to different acceptors) with the hyperthermophilic glycosynthase from Sulfolobus solfataricus acting in dilute sodium formate buffer at pH 4.0 are reported; the use of 4-nitrophenyl beta-glucopyranoside as both donor and acceptor in the self-trans...

In oligosaccharide synthesis the chemo-enzymatic approach is often chosen to obtain the desired glycosidic bond by exploiting the exquisite substrate specificity and stereoselectivity of glycosidases and glycosyltransferases. In recent years the invention of new enzymatic activities, named glycosynthases, created a third route for the enzymatic syn...

Retaining glycosidases promote the hydrolysis of the substrate by following a double-displacement mechanism involving a covalent intermediate. The catalytic residues are a general acid/base catalyst and the nucleophile. Experimental identification of these residues in a specific glycosidase allows for the assigning of the corresponding residues in...

Aphidius ervi is an endophagous braconid, parasitoid of the pea aphid, Acyrthosiphon pisum. A. ervi teratocytes, deriving from the dissociation of the embryonic serosa, synthesize and release two major proteins into the host haemocoel. The gene of one of these proteins has been cloned and characterized. This gene codes for a 15.8 kDa protein belong...

Standard decoding of the genetic information into polypeptides is performed by one of the most sophisticated cell machineries, the translating ribosome, which, by following the genetic code, ensures the correspondence between the mature mRNA and the protein sequence. However, the expression of a minority of genes requires programmed deviations from...