Guillon V

General

Full name : Guillon Virginia

First name : Virginia

Mail : Institut de Recherche Biomedicale des armees - Departement toxicologie et risques chimiques, Bretigny-sur-Orge Cedex

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Country : France

Email : virginia.guillon@irba.fr

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References (4)

Title : Characterization of four BCHE mutations associated with prolonged effect of suxamethonium - Brazzolotto_2021_Pharmacogenomics.J_21_165
Author(s) : Brazzolotto X , Courcelle S , Sauvanet C , Guillon V , Igert A , Kononchik J , Nachon F , Ceppa F , Delacour H
Ref : Pharmacogenomics J , 21(2):165-173 : , 2021
Abstract : Butyrylcholinesterase (BChE) deficiency is characterized by prolonged apnea after the use of muscle relaxants (suxamethonium or mivacurium) in patients who have mutations in the BCHE gene. Here, we report the characterization of four BCHE mutations associated with prolonged effect of suxamethonium (amino acid numbering based on the matured enzyme): p.20delValPheGlyGlyThrValThr, p.Leu88His, p.Ile140del and p.Arg386Cys. Expression of recombinant BCHE mutants, kinetic analysis and molecular dynamics were undertaken to understand how these mutations induce BChE deficiency. Three of the mutations studied (p.20delValPheGlyGlyThrValThr, p.Ile140del and p.Arg386Cys) lead to a "silent" BChE phenotype. Recombinant BCHE expression studies for these mutants revealed BChE activity levels comparable to untransfected cells. Only the last one (hBChE-L88H) presented BChE activity in the transfected cell culture medium. This BChE mutant (p.Leu88His) is associated with a lower k(cat) value compare to the wild-type enzyme. Molecular dynamics simulations analyses suggest that a destabilization of a structure implicated in enzyme activity (-loop) can explain the modification of the kinetic parameter of the mutated protein.
ESTHER : Brazzolotto_2021_Pharmacogenomics.J_21_165
PubMedSearch : Brazzolotto_2021_Pharmacogenomics.J_21_165
PubMedID: 33024248

Title : X-ray structures of human bile-salt activated lipase conjugated to nerve agents surrogates - Touvrey_2019_Toxicology_411_15
Author(s) : Touvrey C , Courageux C , Guillon V , Terreux R , Nachon F , Brazzolotto X
Ref : Toxicology , 411 :15 , 2019
Abstract : The efficiency of human butyrylcholinesterase (BChE) as a stoichiometric bioscavenger of nerve agents is well established. However, wide use is currently limited by production and purification costs. Aiming at identifying an alternative human protein bioscavenger, we looked for an original scaffold candidate by virtual screening of the Protein Data Bank for functional similarity using the "Surfing the Molecules" software (sumo-pbil.ibcp.fr) and a search model based on the BChE active site topology. Besides the expected acetylcholinesterase and butyrylcholinesterase, we identified a set of bile salt activated lipases structures, among which the human pancreatic lipase (hBAL) that shares 34% identity with BChE. We produced the recombinant enzyme in mammalian cells, purified it, and measured the inhibition constants for paraoxon and surrogates of VX, sarin and tabun. We solved the X-ray structure of apo hBAL and conjugates with paraoxon and the surrogates at resolutions in the 2-A range. These structures allow the assessment of hBAL for scavenging nerve agents. They revealed that hBAL has inverted stereoselectivity for the surrogates of nerve agent compared to human cholinesterases. We observed a remarkable flip of the catalytic histidine driven by the chelation of Zn(2+). Dealkylation of the conjugate, aka aging, was solely observed for paraoxon.
ESTHER : Touvrey_2019_Toxicology_411_15
PubMedSearch : Touvrey_2019_Toxicology_411_15
PubMedID: 30359675
Gene_locus related to this paper: human-CEL

Title : Bacterial Expression of Human Butyrylcholinesterase as a Tool for Nerve Agent Bioscavengers Development - Brazzolotto_2017_Molecules_22_
Author(s) : Brazzolotto X , Igert A , Guillon V , Santoni G , Nachon F
Ref : Molecules , 22 : , 2017
Abstract : Human butyrylcholinesterase is a performant stoichiometric bioscavenger of organophosphorous nerve agents. It is either isolated from outdated plasma or functionally expressed in eukaryotic systems. Here, we report the production of active human butyrylcholinesterase in a prokaryotic system after optimization of the primary sequence through the Protein Repair One Stop Shop process, a structure- and sequence-based algorithm for soluble bacterial expression of difficult eukaryotic proteins. The mutant enzyme was purified to homogeneity. Its kinetic parameters with substrate are similar to the endogenous human butyrylcholinesterase or recombinants produced in eukaryotic systems. The isolated protein was prone to crystallize and its 2.5-A X-ray structure revealed an active site gorge region identical to that of previously solved structures. The advantages of this alternate expression system, particularly for the generation of butyrylcholinesterase variants with nerve agent hydrolysis activity, are discussed.
ESTHER : Brazzolotto_2017_Molecules_22_
PubMedSearch : Brazzolotto_2017_Molecules_22_
PubMedID: 29077024
Gene_locus related to this paper: human-BCHE

Title : Serial Femtosecond Crystallography and Ultrafast Absorption Spectroscopy of the Photoswitchable Fluorescent Protein IrisFP - Colletier_2016_J.Phys.Chem.Lett_7_882
Author(s) : Colletier JP , Sliwa M , Gallat FX , Sugahara M , Guillon V , Schiro G , Coquelle N , Woodhouse J , Roux L , Gotthard G , Royant A , Uriarte LM , Ruckebusch C , Joti Y , Byrdin M , Mizohata E , Nango E , Tanaka T , Tono K , Yabashi M , Adam V , Cammarata M , Schlichting I , Bourgeois D , Weik M
Ref : J Phys Chem Lett , 7 :882 , 2016
Abstract : Reversibly photoswitchable fluorescent proteins find growing applications in cell biology, yet mechanistic details, in particular on the ultrafast photochemical time scale, remain unknown. We employed time-resolved pump-probe absorption spectroscopy on the reversibly photoswitchable fluorescent protein IrisFP in solution to study photoswitching from the nonfluorescent (off) to the fluorescent (on) state. Evidence is provided for the existence of several intermediate states on the pico- and microsecond time scales that are attributed to chromophore isomerization and proton transfer, respectively. Kinetic modeling favors a sequential mechanism with the existence of two excited state intermediates with lifetimes of 2 and 15 ps, the second of which controls the photoswitching quantum yield. In order to support that IrisFP is suited for time-resolved experiments aiming at a structural characterization of these ps intermediates, we used serial femtosecond crystallography at an X-ray free electron laser and solved the structure of IrisFP in its on state. Sample consumption was minimized by embedding crystals in mineral grease, in which they remain photoswitchable. Our spectroscopic and structural results pave the way for time-resolved serial femtosecond crystallography aiming at characterizing the structure of ultrafast intermediates in reversibly photoswitchable fluorescent proteins.
ESTHER : Colletier_2016_J.Phys.Chem.Lett_7_882
PubMedSearch : Colletier_2016_J.Phys.Chem.Lett_7_882
PubMedID: 26866390