P Masson

..More studies are needed to strengthen the hypothesis that people with genetic variants of BChE are at higher risk of intoxication from pesticides...

..substrate concentration showed a premature plateau far below V(max). This was due to the loss of substrate availability. The best substrates for rat butyrylcholinesterase were short alkyl homologues, BzCh - BCH4...

..Our results show that OP pesticides and nerve agents bind covalently to human albumin at Tyr411. The presence of Tyr411 on an exposed surface of albumin suggests that an antibody response could be generated against OP-albumin adducts...

..g. cholinergic synaptic clefts. Therefore, physiological or toxicological relevance of the hysteretic behavior and damped oscillations in ChE-catalyzed reactions and inhibition cannot be ruled out...

..Our results revealed HPBP as a new and potent anti HIV molecule that inhibits transcription of the virus, which has not yet been targeted by HAART and therefore opens new strategies in the treatment of HIV infection...

..It is proposed, but not yet proven, that low dose exposure involves OP modification of proteins that have no active site serine...

..Also, BuChE has been integrated in biosensors for detection of organophosphorus compounds and other cholinesterase inhibitors...

..Additional mutations were made for optimizing activity of the new dyad. It is anticipated that these new mutants will have OP hydrolase activity...

..Knowledge of the molecular basis of aging will help to design reactivators of aged ChEs, molecules capable of slowing the aging process, and pseudocatalytic ChE-based bioscavengers...

..The complex inhibition/activation effects of effectors, depending on the integrity of the peripheral anionic site, reflect the allosteric 'cross-talk' between the peripheral anionic site and the catalytic centre...

..Protein engineering can yield enzymes with improved catalytic properties, that are stable during storage and in the bloodstream, and that are immunologically compatible. Large-scale production must be feasible and reasonably inexpensive...

..Interaction of these groups with the ethereal hetero-atom can have a neutral, an additive or an antagonistic effect on transition state stabilization, depending on their molecular structure, size and enantiomeric configuration...

..Therefore, a physiological or toxicological relevance for the hysteresis in BChE cannot be ruled out...

..e., temperature, solvent composition, pH), pressure-induced enzyme unfolding, compressibility changes and pressure-induced rate limiting changes. These different situations are illustrated with several examples...

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..Finally, the cysteines constituting the PON1 disulfide bond (C41 and C352) were essential, but the glycan chains linked to Asn 252 and 323 were not essential for PON1 secretion and activity...

..In mutant enzymes, mutated residues function like check valves: forced penetration of water in the gorge is difficult, release of water is facilitated...

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..It is concluded that double mutations on the PAS induce a conformational change in the active site gorge of butyrylcholinesterase that can alter both substrate binding and enzyme acylation...

..People with the D70G mutation will be resistant to re-activation of their inhibited BuChE by 2-PAM, but this will be offset by the lower rate of irreversible aging of inhibited BuChE, allowing some regeneration by spontaneous hydrolysis...

..Trp residue(s) may be involved in the binding of aromatic substrates...

..Echothiophate and paraoxon were hydrolyzed with the same kcat of 0.75 min-1. This calculates to a rate acceleration of 100,000-fold for hydrolysis of echothiophate and paraoxon by the G117H mutant compared to the nonenzymatic rate...

..Accordingly, CZE is a powerful tool to study protein unfolding/folding quickly and with minimal sample requirements...

..e. forced penetration of water into the gorge is not easily achieved, thereby facilitating the release of water...

..Thus, Km and Ki changes explain the succinyldicholine sensitivity of people carrying the fluoride-2 variant...

..The results indicate that the residue D70 controls the conformational plasticity of the active site gorge of BuChE, and is involved in regulation of the catalytic activity as a function of temperature...

..W430 as well as W231 and W82 appear to form the wall of the active site gorge. Mutation of any of these tryptophans disrupts the architecture of the active site...

..Aromatic/aromatic interactions between the four helical segments are thought to be enhanced by ultrasound-generated pressure...

..Osmolyte-induced change in conformation and/or hydration status of Y332 and other solvent-exposed residues may account for the non-Michaelian behaviour of the D70G mutant...

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..However, we cannot rule out that the effects of mutations on catalytic properties resulted from a remote conformational change and/or misfolding of mutant proteins...

..It was concluded that the active sites for esterase and AAA activities are identical, i.e. S198. This excludes any other residue present in the gorge for being the catalytic nucleophile pole...

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..This study provides the structural basis to design new oximes capable of reactivating phosphoramidyl-hBChE conjugates after intoxication, notably when hBChE is used as pretreatment, or to design BChE-based catalytic bioscavengers...

..The E-->E' transition is accompanied by a negative activation volume (DeltaV++(0)= -45+/-10 ml/mol), and the E' form is more compact than E. Hydration water in the gorge of E' appears to be more structured than in the unprimed form...

..Accordingly, CE can be proposed as a convenient tool to study protein stability and denaturation processes...

..However, designed mutants of this enzyme capable of degrading organophosphates have to be engineered to enhance their thermostability...

..3. The future prospects of affinity electrophoresis, e.g., investigations on the esteratic site and exploration of the carbohydrate moiety, are emphasized in the concluding section...

..A first-order deactivation model describing the kinetics of the remnant inactivation of BuChE is proposed...

..Pesticides and certain stereoisomers of nerve agents are expected to undergo aging by breaking the P-O bond...

..1H-NMR measurements provided evidence for a slow equilibrium between two BzCh conformers. Binding of the conformationally preferred substrate conformer leads to products...

..In our research group, a directed evolution project has been initiated on a bacterial phosphotriesterase, a promising enzyme, capable of efficiently detoxifying organophosphorus nerve agents...

..Fluorescence of PON1 complexed to terbium ion shows that at least one tryptophan is close to the calcium binding site...

..Nature of the organ(s) and mechanisms involved in enzyme release will need further investigations as it may represent a mechanism of defence, i.e. VX scavenging, that could advantageously be exploited...

..Determination of the PON1 three-dimensional structure is a key step toward improvement of the enzyme functional properties...

..5 degrees C). We attempted to detect these two transitions by differential scanning calorimetry, UV spectrophotometry and circular dichroism measurements. These transitions have been ascribed to subtle cold-induced conformational changes...

..e., the classical X-R scission pathway. In contrast, isomalathion aged through both X-R and P-X pathways; the main aged product resulted from P-S bond cleavage and a minor product resulted from O-C and/or S-C bond cleavage...

..We suggest that a similar mechanism is also possible for oxime reactivation of phosphylated cholinesterases. This opens new perspectives in terms of reactivator design...

..6-1 mM), albumin may participate in hydrolysis of aryl acylamides through second-order kinetics. This suggests that albumin may have a role in the metabolism of endogenous and exogenous aromatic amides, including drugs and xenobiotics...

..The finding that AChE is activated by excess substrate supports the idea that binding of a second substrate molecule to the PAS induces a conformational change that reorganizes the active site...

..It is concluded that OP bind covalently to tubulin, and that this binding could explain cognitive impairment associated with OP exposure...

..Acetylated lysines formed adducts that were even more stable. In conclusion, the pseudo-esterase activity of albumin is the result of irreversible acetylation of 82 residues and is not the result of turnover...

..The reported 3D data will help in the design of new oximes capable of reactivating tabun-ChE conjugates...

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..The method uses 1 ml of serum and detects low-level exposure associated with as little as 20% inhibition of plasma butyrylcholinesterase...

..Modified peptides are difficult to find when the OP bears no radiolabel and no tag. The characteristic MSMS fragment ions are valuable because they are identifiers for OP-tyrosine, independent of the peptide...

..coli and natural human PON1 provide information for the future rational design of stable mutants of PON1-based catalytic scavengers to be used as safe and effective countermeasures to OP intoxication...

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..At higher pressures, the enzyme was converted to forms which were irreversibly inactive and had a higher apparent molecular mass, suggesting aggregation or denaturation...

..Maintaining a stable environment for such administrable human rPON1 should, at least, preserve the anti-atherogenic activity of the enzyme...

..73 A3 per Da (estimated 60% solvent) for a single molecule of recombinant BChE in the asymmetric unit. The crystal structure of butyrylcholinesterase will help elucidate unsolved issues concerning cholinesterase mechanisms in general...

..We propose that the proline-rich peptides organize the 4 subunits of BChE into a 340 kDa tetramer, by interacting with the C-terminal BChE tetramerization domain...

..Since human plasma contains no carboxylesterase, only BChE, PON1, and albumin esterases need to be considered when evaluating hydrolysis of an ester drug in human plasma...

..5-67 degrees C, 58-64 degrees C and 53-64 degrees C respectively). Requirements for optimum activity of PTE (displayed by Co2+-PTE) and maximum stability (displayed by Zn2+-PTE) were demonstrated...

..Diethoxyphosphate-labeled Tyr 411 was stable for months at pH 7.4. These results will be useful in the development of specific antibodies to detect OP exposure and to engineer albumin for use as an OP scavenger...

..The extent to which tyrosine modification by OP can occur in vivo and the toxicological implications of such modifications require further investigation...

..A hybrid recombinant PON1 was shown more thermostable than the human enzyme, and its stability was unaffected by the presence of HPBP. Altogether, the results strongly encourage further study of the human enzyme...

..In conclusion, pressure and thermal stability of mutants provided evidence that the residue E202 is a determinant of structural and functional stability of HuAChE...

..Soman-albumin adducts detected by mass spectrometry could be useful for the diagnosis of soman exposure...

..The mechanism explaining the catalytic behaviour of butyrylcholinesterase at low pH in the presence of excess substrate remains to be elucidated...

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..Both wild-type and G117H BChE were in the epithelial cells of the villi. These assays can be used to identify OP-resistant cholinesterases in culture medium and in animal tissues...

..Finally, capillary electrophoresis has proved to be a pertinent tool to validate the conformity of purified enzymes to a status of biopharmaceutical...

..Moreover, high pressure can be used instead of heart or chemicals for sterilization...

..Issues addressed in this work are important for future biotechnological developments of PTE as a detoxifying enzyme...

..Because inhibitors used in purification of ChEs by affinity chromatography are charged, CE remains a leading method to estimate enzyme stability and detect the presence of bound hidden ligands...

..Identification of other OP-modified proteins may lead to an understanding of neurotoxic symptoms that appear long after the initial OP exposure...

..The 29 residue active site tryptic peptide labeled with the nerve agents soman or sarin was identified...

..Changes of HDL size and shape could strongly affect the binding affinity and stability of HuPON1 and result in reduced antioxidative capacity of the lipoprotein...

..Finally, CZE was proved to be a pertinent tool to validate the conformity of purified enzymes to a status of biopharmaceutical...

..Thermal stability, susceptibility to high electric field, alteration of stability by bound ligands and the role of associated cations in metalloenzymes have been successfully investigated...

..Relevance of these data to understand in vivo physiological PON1 functioning is mandatory...

..1 MPa) and 75 degrees C. The use of high pressure for spore inactivation represents a considerable improvement over other available methods of spore inactivation and could be of interest for antigenic spore preparation...

..Hence, for decontamination purposes under extreme Earth temperatures, wild type and engineered mutants of PTE substituted with other metal cations should be evaluated...

..Altogether, results underline the importance of the OP nitrogen atom at beta-position and of its substituents, even though the intimate mechanism of laccase-catalyzed degradation is not yet known...

..DFP was characterised as a slow-binding substrate. This mutant is the first mutant of AChE capable of hydrolysing organophosphates. However, the overall OPase efficiency was greatly decreased compared to G117H butyrylcholinesterase...

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..At the concentrations around 40 mm, well within the range of TEA competition at both substrate binding sites, it lowers the activity of all tested enzymes...

..This example demonstrates that promiscuous activities probably constitute a large and efficient reservoir for the creation of novel catalytic activities...

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..Hence, the catalytic power of cholinesterases can largely be understood in terms of their ability to stabilize tetrahedral intermediates in the multistep reaction mechanism...

..This structural characterization of acetylcholinesterase 'at work' is consistent with the idea that choline exits from the enzyme after catalysis either via the gorge or via an alternative 'backdoor' trajectory...

..Kinetic assays were carried out with paraoxon as organophosphate model. Three regioisomers hydrolyzed paraoxon, although the paraoxon-leaving group, para-nitrophenol, was not released from the beta-cyclodextrin torus...

..An electron density peak close to the catalytic Ser(198) has been modeled as bound butyrate...

..The S13 subunit may possess multiple functions, including a deubiquitinylating activity and distinct activities essential for cell cycle progression that require the conserved C120 residue...

..Here, the crystallization conditions and data collection for the hyperthermophilic S. solfataricus PTE are reported...