Resolving pathways of interaction of mipafox and a sarin analog with human acetylcholinesterase by kinetics, mass spectrometry and molecular modeling approaches

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F62690094%3A18450%2F16%3A50004656" target="_blank" >RIV/62690094:18450/16:50004656 - isvavai.cz</a>

Výsledek na webu

<a href="http://dx.doi.org/10.1007/s00204-015-1481-1" target="_blank" >http://dx.doi.org/10.1007/s00204-015-1481-1</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1007/s00204-015-1481-1" target="_blank" >10.1007/s00204-015-1481-1</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Resolving pathways of interaction of mipafox and a sarin analog with human acetylcholinesterase by kinetics, mass spectrometry and molecular modeling approaches

Popis výsledku v původním jazyce

Inhibited AChE can be reactivated by cleavage of the Ser–phosphorus bond either spontaneously or through a reaction with nucleophilic agents, such as oximes. At the same time, the inhibited AChE adduct can lose part of the molecule by progressive dealkylation over time in a process called aging. Reactivation of the aged enzyme has not yet been demonstrated. Here, our goal was to study oxime reactivation and aging reactions of human AChE inhibited by mipafox or a sarin analog (Flu-MPs, fluorescent methylphosphonate). Progressive reactivation was observed after Flu-MPs inhibition using oxime 2-PAM. However, no reactivation was observed after mipafox inhibition with 2-PAM or the more potent oximes used. A peptide fingerprinted mass spectrometry (MS) method, which clearly distinguished the peptide with the active serine (active center peptide, ACP) of the human AChE adducted with OPs, was developed by MALDITOF and MALDI-TOF/TOF. The ACP was detected with a diethyl-phosphorylated adduct after paraoxon inhibition, and with an isopropylmethyl-phosphonylated and a methyl-phosphonylated adduct after Flu-MPs inhibition and subsequent aging. Nevertheless, nonaged nonreactivated complexes were seen after mipafox inhibition and incubation with oximes, where MS data showed an ACP with an NN diisopropyl phosphoryl adduct. The kinetic experiments howed no reactivation of activity. The computational molecular model analysis of the mipafox-inhibited hAChE plots of energy versus distance between the atoms separated by dealkylation showed a high energy demand, thus little aging probability. However, with Flu-MPs and DFP, where aging was observed in our MS data and in previously published crystal structures, the energy demand calculated in modeling was lower and, consequently, aging appeared as a more likely reaction. We document here direct evidence for a phosphorylated hAChE refractory to oxime reactivation, although we observed no aging.

Název v anglickém jazyce

Resolving pathways of interaction of mipafox and a sarin analog with human acetylcholinesterase by kinetics, mass spectrometry and molecular modeling approaches

Popis výsledku anglicky

Inhibited AChE can be reactivated by cleavage of the Ser–phosphorus bond either spontaneously or through a reaction with nucleophilic agents, such as oximes. At the same time, the inhibited AChE adduct can lose part of the molecule by progressive dealkylation over time in a process called aging. Reactivation of the aged enzyme has not yet been demonstrated. Here, our goal was to study oxime reactivation and aging reactions of human AChE inhibited by mipafox or a sarin analog (Flu-MPs, fluorescent methylphosphonate). Progressive reactivation was observed after Flu-MPs inhibition using oxime 2-PAM. However, no reactivation was observed after mipafox inhibition with 2-PAM or the more potent oximes used. A peptide fingerprinted mass spectrometry (MS) method, which clearly distinguished the peptide with the active serine (active center peptide, ACP) of the human AChE adducted with OPs, was developed by MALDITOF and MALDI-TOF/TOF. The ACP was detected with a diethyl-phosphorylated adduct after paraoxon inhibition, and with an isopropylmethyl-phosphonylated and a methyl-phosphonylated adduct after Flu-MPs inhibition and subsequent aging. Nevertheless, nonaged nonreactivated complexes were seen after mipafox inhibition and incubation with oximes, where MS data showed an ACP with an NN diisopropyl phosphoryl adduct. The kinetic experiments howed no reactivation of activity. The computational molecular model analysis of the mipafox-inhibited hAChE plots of energy versus distance between the atoms separated by dealkylation showed a high energy demand, thus little aging probability. However, with Flu-MPs and DFP, where aging was observed in our MS data and in previously published crystal structures, the energy demand calculated in modeling was lower and, consequently, aging appeared as a more likely reaction. We document here direct evidence for a phosphorylated hAChE refractory to oxime reactivation, although we observed no aging.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10609 - Biochemical research methods

Projekt

—

Návaznosti

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2016

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Archives of toxicology

ISSN

0340-5761

e-ISSN

—

Svazek periodika

90

Číslo periodika v rámci svazku

3

Stát vydavatele periodika

DE - Spolková republika Německo

Počet stran výsledku

14

Strana od-do

603-616

Kód UT WoS článku

000370343000009

EID výsledku v databázi Scopus

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