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These studies are focused on antagonizing organophosphorous (OP) intoxications by a new conceptual approach using recombinant enzymes encapsulated within sterically stabilized liposomes to enhance diisopropylfluorophosphate (DFP) degradation. The OP hydrolyzing enzyme, organophosphorous acid anhydrolase (OPAA), encapsulated within the liposomes, wa...
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... to DFP, and against soman, should approximate that of DFP. Enzyme kinetic data indicate that paraoxon has a high affinity for OP hydrolase (OPH), but this enzyme is less active for DFP, sarin, and soman (Table 2). On extrapolations from these enzyme kinetic data, OPAA seems to be a more appropriate enzyme for studies on DFP, sarin, and soman ( Fig. 1 and Table ...
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Subchronic pretreatment with physostigmine (PHY) (0.0125 mg/kg/h) leading to a blood acetylcholinesterase inhibition of about 30% caused no side effects when applied to marmoset monkeys. This was evident on behavioral parameters and on EEG and cortical visual evoked response. Furthermore, this treatment regime, followed by atropine as postintoxicat...
The synthesis, characterization and sensing features of a novel probe 1 for the selective chromogenic recognition of diisopropylfluorophosphate (DFP), a sarin and soman mimic, in 99:1 (v/v) water/acetonitrile and in the gas phase is reported. Colour modulation is based on the combined reaction of phosphorylation of 1 and fluoride-induced hydrolysis...
Citations
... Further study suggested that Alteromonas haloplanktis OPAA is a type of prolidase. Alteromonas prolidase can hydrolyze G-type nerve agents and DFP, soman, sarin, and cyanide containing tabun, therefore is used as an antidote for poisoning (Cheng et al., 1997;Cheng et al., 1999;Petrikovics et al., 2000). Native and recombinant human prolidase also catalyzes these nerve agents with A252R or P365R substitution mutation of the enzyme, improving the catalytic activity against organophosphorus compounds (Chandrasekaran et al., 2013;Yun et al., 2017). ...
Prolidase (peptidase D), encoded by the PEPD gene, is a ubiquitously expressed cytosolic metalloproteinase, the only enzyme capable of cleaving imidodipeptides containing C-terminal proline or hydroxyproline. Prolidase catalyzes the rate-limiting step during collagen recycling and is essential in protein metabolism, collagen turnover, and matrix remodeling. Prolidase, therefore plays a crucial role in several physiological processes such as wound healing, inflammation, angiogenesis, cell proliferation, and carcinogenesis. Accordingly, mutations leading to loss of prolidase catalytic activity result in prolidase deficiency a rare autosomal recessive metabolic disorder characterized by defective wound healing. In addition, alterations in prolidase enzyme activity have been documented in numerous pathological conditions, making prolidase a useful biochemical marker to measure disease severity. Furthermore, recent studies underscore the importance of a non-enzymatic role of prolidase in cell regulation and infectious disease. This review aims to provide comprehensive information on prolidase, from its discovery to its role in health and disease, while addressing the current knowledge gaps.
... In addition, the use of liposomal systems does not require blood group phenotyping. The first works of Petrikovics's group using sterically stabilized liposomes as a carrier system were for encapsulation of recombinant PTE [140], OPAA [141,142], recombinant OPH [143]. OPs easily penetrate into liposomes circulating in the bloodstream, and by-products leave liposomes after enzymatic hydrolysis. ...
The introduction of enzyme nanoreactors in medicine is relatively new. However, this technology has already been experimentally successful in cancer treatments, struggle against toxicity of reactive oxygen species in inflammatory processes, detoxification of drugs and xenobiotics, and correction of metabolic and genetic defects by using encapsulated enzymes, acting in single or cascade reactions. Biomolecules, e.g. enzymes, antibodies, reactive proteins capable of inactivating toxicants in the body are called bioscavengers.
In this review, we focus on enzyme-containing nanoreactors for in vivo detoxification of organophosphorous compounds (OP) to be used for prophylaxis and post-exposure treatment of OP poisoning. A particular attention is devoted to bioscavenger-containing injectable nanoreactors operating in the bloodstream. The nanoreactor concept implements single or multiple enzymes and cofactors co-encapsulated in polymeric semi-permeable nanocontainers. Thus, the detoxification processes take place in a confined space containing highly concentrated bioscavengers. The article deals with historical and theoretical backgrounds about enzymatic detoxification of OPs in nanoreactors, nanoreactor polymeric enveloppes, realizations and advantages over other approaches using bioscavengers.
... A representative comparison table of OPAA-FL with P-F and P-O bond containing OPs, and OPAA WT with P-O bond containing compounds, as well as a comparison of OPAA FL and wild-type OPAA for the hydrolysis of P-S bond containing Russian VX is given in Tables S3 and S4, respectively. A group of scientists in the year 2000 proved that by encapsulating OPAA hydrolyzing enzyme in conjugation with 2-PAM and atropine in lysosome to provide 23 LD50s of protection against DFP (Petrikovics et al. 2000). Our report attempts to evaluate the OPAA-FL variant as a potential candidate for the enzymatic hydrolysis of paraoxon, and it is expected that the studies would aid efforts towards developing effective sensors and/or mutant enzymes for the purpose of detection and/or degradation of organophosphates, especially ones with the P-O bond such as paraoxon. ...
Indiscriminate use of organophosphorus (OP)-based insecticides is a great concern to human health because of bioaccumulation-induced health hazards. Potentially fatal consequences and limited treatment methods of OP poisoning necessitate the need for the development of reliable, selective, cost-effective, and sensitive methods of OP detection. To tackle this issue, the development of effective devices and methods is required to sensitively detect as well as degrade OPs. Enzymatic sensor systems have gained popularity due to high catalytic activity, enhanced detection limits, and high sensitivity with the environmentally benign operation. Organophosphorus acid anhydrolase (OPAA) from Alteromonas sp. JD6.5 is capable of hydrolyzing the P-F, P-O, P-S, and P-CN bonds, in OPs, including nerve agents of the G/V-series. Several mutants of OPAA are reported which have greater activity against various OPs. In this study, recombinant expression of the OPAA-FL variant in Escherichia coli was performed, purified, and subsequently tested for activity against ethyl paraoxon. OPAA-FL variant showed its optimum activity at pH 8.5 and 50 °C. Colorimetric and fluorometric assays were used for estimation of ethyl paraoxon based on p-nitrophenol and fluorescein isothiocyanate (FITC) fluorescence intensity, respectively. Colorimetric and fluorometric assay estimation indicates that ethyl paraoxon can be estimated in the linear range of 0.01 to 1 mM and 0.1 to 0.5 mM, with LOD values 0.04 mM and 0.056 mM, respectively. Furthermore, the OPAA-FL variant was immobilized into alginate microspheres for colorimetric detection of ethyl paraoxon and displayed a linear range of 0.025 to 1 mM with a LOD value of 0.06 mM. • Biosensing of paraoxon with purified and encapsulated OPAA-FL variant. • Colorimetric and fluorometric biosensing assay developed using OPAA-FL variant for paraoxon. • First report on alginate encapsulation of OPAA-FL variant for biosensing of paraoxon. Graphical abstract
... The authors showed that the encapsulated enzyme was able to significantly out-perform the free enzyme in both plasma and controls in buffered solutions. Liposomal encapsulation has also been used for the colocalization of enzyme and other therapeutics such as atropine as described for OPAA by Petrikovics (Petrikovics et al., 2000). These studies suggest that encapsulation of enzyme could be used as pathways for in vivo delivery of detoxifying therapeutics, or as discussed below, vehicles for field deployment. ...
Organophosphate compounds are ubiquitously employed as agricultural pesticides and maintained as chemical warfare agents by several nations. These compounds are highly toxic, show environmental persistence and accumulation, and contribute to numerous cases of poisoning and death each year. While their use as weapons of mass destruction is rare, these never fully disappear into obscurity as they continue to be tools of fear and control by governments and terrorist organizations. Beyond weaponization, their wide-scale dissemination as agricultural products has led to environmental accumulation and intoxication of soil and water across the globe. Therefore, there is a dire need for rapid and safe agents for environmental bioremediation, personal decontamination, and as therapeutic detoxicants. Organophosphate hydrolyzing enzymes are emerging as appealing targets to satisfy decontamination needs owing to their ability to hydrolyze both pesticides and nerve agents using biologically-derived materials safe for both the environment and the individual. As the release of genetically modified organisms is not widely accepted practice, researchers are exploring alternative strategies of organophosphate bioremediation that focus on cell-free enzyme systems. In this review, we first discuss several of the more prevalent organophosphorus hydrolyzing enzymes along with research and engineering efforts that have led to an enhancement in their activity, substrate tolerance, and stability. In the later half we focus on advances achieved through research focusing on enhancing the catalytic activity and stability of phosphotriesterase, a model organophosphate hydrolase, using various approaches such as nanoparticle display, DNA scaffolding, and outer membrane vesicle encapsulation.
... La pharmacocinétique et l'immunogénicité notamment doivent être évaluées afin de limiter la quantité d'enzyme injectée pour limiter le risque d'une réaction du système immunitaire [68,120]. Dans cette optique, la PEGylation [121], les liposomes [122,123] et l'encapsulation dans d'autres nano-conteneurs [124,125] sont d'un intérêt capital. Bien que nécessitant de plus amples études pour des applications en prophylaxie et en traitement, les enzymes sont extrêmement intéressantes pour la décontamination externe comme alternative aux méthodes chimiques et physiques conventionnelles telles que l'hypochlorite, l'hydroxyde de sodium, la terre à Foulon ou le RSDL (Reactive Skin Decontamination Lotion) [33]. ...
Résumé
Les organophosphorés (OPs) sont des composés chimiques toxiques développés en tant qu’insecticides et agents de guerre. Ils sont le plus souvent neurotoxiques et perturbent le système nerveux en bloquant la transmission cholinergique. Les OPs seraient responsables de nombreuses intoxications dans le monde. La toxicité de ces composés peut être liée soit à une exposition aigüe, soit à une exposition chronique et leurs effets ont été étudiés sur différents modèles animaux. Ces derniers ont également été utilisés pour évaluer l’efficacité d’antidotes. Des stratégies basées sur l’utilisation d’enzymes pouvant piéger (bioscavengers stœchiométriques) ou dégrader (bioscavengers catalytiques) les OPs, ont été notamment étudiées puisqu’elles permettent une décontamination efficace, sans toxicité, ni impact environnemental. Cette revue rassemble les résultats obtenus in vivo avec des enzymes à travers trois niveaux de prise en charge: la prophylaxie, le traitement et la décontamination externe. L’efficacité des traitements enzymatiques évalués dans différents modèles animaux, est présentée et la pertinence de ces modèles est également discutée pour une meilleure extrapolation à l’homme.
... However, the catalytic efficiencies k cat /K m of OPH towards fluoride-containing nerve agents including DFP are ranging from 1.0 × 10 4 to 8.0 × 10 4 M -1 s -1 , much lower than that towards its optimal substrate ethyl-paraoxon (k cat /K m = 5.5 × 10 7 M -1 s -1 ) [8,11,39,43]. To date, the desirable enzyme(s) to degrade DFP has been recorded as squid diisopropylfluorophosphatase (DFPase; EC 3.1.8.2) with k cat /K m 1.4 × 10 5 M −1 s −1 [11,13,39,40,41] and Alteromonas organophosphorus acid anhydrolases (OPAA; EC 3.1.8.2) with about one order of magnitude higher k cat /K m (5.5 × 10 5 M −1 s −1 ) [44][45][46][47]. In the present study, the optimal OPH activity towards DFP has been assessed by k cat /K m 541.7 ± 12.6 mM -1 s -1 (approximately 5.4 × 10 5 M -1 s -1 ) at 300 mM TEA condition (Table 6), desirably higher than DFPase and reaching at the level of OPAA, the best DFP-degrading enzyme ever reported. ...
... In the present study, the optimal OPH activity towards DFP has been assessed by k cat /K m 541.7 ± 12.6 mM -1 s -1 (approximately 5.4 × 10 5 M -1 s -1 ) at 300 mM TEA condition (Table 6), desirably higher than DFPase and reaching at the level of OPAA, the best DFP-degrading enzyme ever reported. OPAA has been well exploited to detoxify fluoride-containing OPs and successfully encapsulated as dentritic-enzyme complexes with pralidoxime and/or atropine to protect AChE and/or cholinesterase against DFP [47][48][49]. In contrast, despite of having a wide OPs spectrum, OPH has been not so good to degrade organophosphofluoridates including DFP, somewhat reducing the practical value of this enzyme. ...
Aminoalcohols have been addressed as activating buffers for alkaline phosphatase. However, there is no record on the buffer activation regarding organophosphorus hydrolase (OPH). Here we reported the activating effects of aminoalcohols on OPH-catalyzed hydrolysis of diisopropylfluorophosphate (DFP), an analog molecule of G-type warfare agents. The kinetic parametors kcat, Vmax and kcat/Km in the OPH reaction were remarkably increased in the buffers (pH 8.0, 25°C) containing aminoalcohols with C2 between nitrogen (N) and oxygen (O) in their structures, including triethanolamine (TEA), diethanolamine, monoethanolamine, 1-amino-2-propanol, 2-amino-2-methyl-1-propanol, and triisopropanolamine. In contrast, much lower or no rate-enhancing effects were observed in the adding of amines, alcohols, amine/alcohol mixtures, or 3-amino-1-propanol (C3 between N and O). The 300 mM TEA further increased DFP-degrading activities of OPH mutants F132Y and L140Y, the previously reported OPH mutants with desirable activities towards DFP. However, the treatment of ethylenediaminetetraacetate (EDTA) markedly abolished the TEA-induced activation of OPH. The product fluoride effectively inhibited OPH-catalyzed hydrolysis of DFP by a linear mixed inhibition (inhibition constant Ki ~ 3.21 mM), which was partially released by TEA adding at initial or later reaction stage. The obtained results indicate the activation of OPH by aminoalcohol buffers could be attributed to the reduction of fluoride inhibition, which would be beneficial to the hydrolase-based detoxification of organophosphofluoridate.
... 8,9 Previously, biodegradable liposome nanocarriers have been shown to be effective at providing functionally significant amounts of highly purified enzymes in the bloodstream while protecting the enzymes for over 2 days. 10 Highly desirable, nevertheless, are approaches that may yield comparably high activity and stability, while providing ancillary benefits such as simpler handling and storage of enzymes. 11 Metal−organic frameworks (MOFs), 12 consisting of metal ions/clusters and coordinated organic linkers, represent a class of highly crystalline porous materials with regular crystal morphology, adjustable pore diameters, and tunable function-alities. ...
We report the synthesis and characterization of a water-stable zirconium metal–organic framework (MOF), NU-1003, featuring the largest mesoporous aperture known for a zirconium MOF. This material has been used to immobilize the nerve agent hydrolyzing enzyme, organophosphorus acid anhydrolase (OPAA). The catalytic efficiency of immobilized OPAA in nanosized NU-1003 is significantly increased compared to that of OPAA immobilized in microsized NU-1003 and even exceeds that of the free OPAA enzyme. This paper highlights a method for rapid and highly efficient hydrolysis of nerve agents using nanosized enzyme carriers.
... Natural molecules can permeate vesicle membranes more easily than their charged forms, and if a vesicle supports a pH gradient (acidic or basic for weak bases or acids, respectively), the unionized compound diffuses into such a vesicle where it is ionized and trapped 128 (Fig. 23b). These principles have been used in liposomes encapsulating, for example, sulfurtransferase enzyme degrading cyanide, 129 or organophosphorus acid anhydrolase degrading organophosphorus agents, 130,131 and in liposomes whose pH = 4 interior captured the anticancer drug doxorubicin, decreasing its toxicity while maintaining the drug's anti-tumor potency. 132 Selective membrane permeability and maintenance of concentration differences were also used to modulate reactivity of peptides. ...
Under non-equilibrium conditions, liquid droplets coupled to their environment by sustained flows of matter and/or energy can become “active” systems capable of various life-like functions. When “fueled” by even simple chemical reactions, such droplets can become tactic and can perform “intelligent” tasks such as maze solving. With more complex chemistries, droplets can support basic forms of metabolism, grow, self-replicate, and exhibit evolutionary changes akin to biological cells. There are also first exciting examples of active droplets connected into larger, tissue-like systems supporting droplet-to-droplet communication, and giving rise to collective material properties. As practical applications of droplets also begin to appear (e.g., in single-cell diagnostics, new methods of electricity generation, optofluidics, or sensors), it appears timely to review and systematize progress in this highly interdisciplinary area of chemical research, and also think about the avenues (and the roadblocks) for future work.
... A schematic illustration of the three major types of enzyme-nanocarrier architectures for detoxification through intravenous delivery. [75][76][77][78]. Because their ease of preparation and excellent biocompatibility, liposomes have been extensively explored as delivery carriers for many therapeutic enzymes; however, despite the improved circulating time and therapeutic effects, the delivery efficiency is still far from optimal. ...
... Preclinical study [75][76][77]199,200] ...
... As an artificial replacement of erythrocytes, liposomal carriers were also employed for the delivery of phosphotriesterases. Several different phosphotriesterases, including OPAA [75,77], OPH [76,199], and paraoxonase-1 (PON1) [200], were stabilized by liposomal encapsulation, resulting in extended circulating half-life and enhanced protection against the lethal effect of paraoxon. However, several reports indicated that the activity of encapsulated enzymes appeared a first order kinetics, suggesting that the diffusion limitation of OP compounds caused by the liposome may weaken the detoxification effects of these enzymes. ...
Life relies on numerous biochemical processes working synergistically and correctly. Certain substances disrupt these processes, inducing living organism into an abnormal state termed intoxication. Managing intoxication usually requires interventions, which is referred as detoxification. Decades of development on detoxification reveals the potential of enzymes as ideal therapeutics and antidotes, because their high substrate specificity and catalytic efficiency are essential for clearing intoxicating substances without adverse effects. However, intrinsic shortcomings of enzymes including low stability and high immunogenicity are major hurdles, which could be overcome by delivering enzymes with specially designed nanocarriers. Extensive investigations on protein delivery indicate three types of enzyme-nanocarrier architectures that show more promise than others for systemic detoxification, including liposome-wrapped enzymes, polymer-enzyme conjugates, and polymer-encapsulated enzymes. This review highlights recent advances in these nano-architectures and discusses their applications in systemic detoxifications. Therapeutic potential of various enzymes as well as associated challenges in achieving effective delivery of therapeutic enzymes will also be discussed.
Copyright © 2015. Published by Elsevier B.V.
... The enzyme engineering strategies are applied to increase the strength and stability such as enzyme capsulation [109,110]; changes in affinity of enzyme [111], changes in the specificity of the enzyme [112], a direct enzyme conjugation with quantum dots CDS [113], enzyme immobilization on nano-porous silica substrate [114,115]; cell surface display [116], and secretory enzyme expression [117]. ...
Daily, organophosphorus compounds (OPs) in human life, has found wide
applications. Although OPs have biodegradability potential, they induce clinical
problems in humans and other organism. Different methods are used to detoxify
these compounds. In the meantime, biodegradation is preferred as a compatible
way to the environment since it produces less toxic compounds. Enzymes capable
to degrade the OPs are of the most important items in the biodegradation. Genetic
manipulation involved in the production of these enzymes has been employed in
bacteria, and finally, is used for the mass production of recombinant
microorganisms. In this paper, the role of organophosphates on human life and the
ways to destroy toxic organophosphates are studied.
