Mass spectrometry based proteomic approach for the screening of butyrylcholinesterase adduct formation with organophosphates

Yaroslav Dubrovskii, Ekaterina Murashko, Olga Chuprina, Petr Beltyukov, Andrey Radilov, Nikolay Solovyev, Vladimir Babakov

Research output

2 Citations (Scopus)

Abstract

Organophosphates’ toxic effect causes covalent binding to serine-198 in the active site of human plasma butyrylcholinesterase (BChE) with loss of enzymatic function (covalent inhibition). Mass spectrometric detection of modified FGESAGAAS peptide at the active site is a powerful exposure biomarker tool. The aim of this study was to develop mass spectrometry-based method for BChE adduct formation screening, avoiding the use of standard peptides. Immunomagnetic separation of proteins from plasma was optimized. Commercially available anti-butyrylcholinesterase monoclonal antibodies, immobilized on magnetic beads, resulted in stable and reusable affinity sorbent. The method was tested on horse serum BChE and real human plasma from healthy donors, treated with Russian VX (VR). The BChE isolated from blood plasma was digested with pepsin and analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS). The method was evaluated by using synthetic peptides and by comparison to the enzymatic activity Ellman's assay. The minimum concentration of VR exposure, resulting in detectable VR-adduct, was 0.2 ng/mL, which corresponded to the relative BChE inhibition of less than 2%. Adduct formation assessment was performed via monitoring of decrease in non-modified peptide LC-MS/MS signal and increase in VR-modified peptide signal. The designed approach was tested in a pilot study with 5 blood samples from healthy volunteers. Mass spectrometry-based method for BChE adduct formation was found to be in agreement with Ellman's inhibition assay, so the method is applicable for direct BChE inhibition assessment.

Original languageEnglish
Pages (from-to)374-382
Number of pages9
JournalTalanta
Volume197
DOIs
Publication statusPublished - 15 May 2019

Fingerprint

Butyrylcholinesterase
Organophosphates
Proteomics
Mass spectrometry
Mass Spectrometry
Screening
Plasma (human)
Peptides
Assays
Catalytic Domain
Blood
Immunomagnetic Separation
Poisons
Pepsin A
Liquid chromatography
Biomarkers
Protein Sorting Signals
Tandem Mass Spectrometry
Sorbents
Liquid Chromatography

Scopus subject areas

  • Analytical Chemistry
  • Chemistry(all)
  • Biochemistry
  • Spectroscopy

Cite this

Dubrovskii, Yaroslav ; Murashko, Ekaterina ; Chuprina, Olga ; Beltyukov, Petr ; Radilov, Andrey ; Solovyev, Nikolay ; Babakov, Vladimir. / Mass spectrometry based proteomic approach for the screening of butyrylcholinesterase adduct formation with organophosphates. In: Talanta. 2019 ; Vol. 197. pp. 374-382.
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title = "Mass spectrometry based proteomic approach for the screening of butyrylcholinesterase adduct formation with organophosphates",
abstract = "Organophosphates’ toxic effect causes covalent binding to serine-198 in the active site of human plasma butyrylcholinesterase (BChE) with loss of enzymatic function (covalent inhibition). Mass spectrometric detection of modified FGESAGAAS peptide at the active site is a powerful exposure biomarker tool. The aim of this study was to develop mass spectrometry-based method for BChE adduct formation screening, avoiding the use of standard peptides. Immunomagnetic separation of proteins from plasma was optimized. Commercially available anti-butyrylcholinesterase monoclonal antibodies, immobilized on magnetic beads, resulted in stable and reusable affinity sorbent. The method was tested on horse serum BChE and real human plasma from healthy donors, treated with Russian VX (VR). The BChE isolated from blood plasma was digested with pepsin and analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS). The method was evaluated by using synthetic peptides and by comparison to the enzymatic activity Ellman's assay. The minimum concentration of VR exposure, resulting in detectable VR-adduct, was 0.2 ng/mL, which corresponded to the relative BChE inhibition of less than 2{\%}. Adduct formation assessment was performed via monitoring of decrease in non-modified peptide LC-MS/MS signal and increase in VR-modified peptide signal. The designed approach was tested in a pilot study with 5 blood samples from healthy volunteers. Mass spectrometry-based method for BChE adduct formation was found to be in agreement with Ellman's inhibition assay, so the method is applicable for direct BChE inhibition assessment.",
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Mass spectrometry based proteomic approach for the screening of butyrylcholinesterase adduct formation with organophosphates. / Dubrovskii, Yaroslav; Murashko, Ekaterina; Chuprina, Olga; Beltyukov, Petr; Radilov, Andrey; Solovyev, Nikolay; Babakov, Vladimir.

In: Talanta, Vol. 197, 15.05.2019, p. 374-382.

Research output

TY - JOUR

T1 - Mass spectrometry based proteomic approach for the screening of butyrylcholinesterase adduct formation with organophosphates

AU - Dubrovskii, Yaroslav

AU - Murashko, Ekaterina

AU - Chuprina, Olga

AU - Beltyukov, Petr

AU - Radilov, Andrey

AU - Solovyev, Nikolay

AU - Babakov, Vladimir

PY - 2019/5/15

Y1 - 2019/5/15

N2 - Organophosphates’ toxic effect causes covalent binding to serine-198 in the active site of human plasma butyrylcholinesterase (BChE) with loss of enzymatic function (covalent inhibition). Mass spectrometric detection of modified FGESAGAAS peptide at the active site is a powerful exposure biomarker tool. The aim of this study was to develop mass spectrometry-based method for BChE adduct formation screening, avoiding the use of standard peptides. Immunomagnetic separation of proteins from plasma was optimized. Commercially available anti-butyrylcholinesterase monoclonal antibodies, immobilized on magnetic beads, resulted in stable and reusable affinity sorbent. The method was tested on horse serum BChE and real human plasma from healthy donors, treated with Russian VX (VR). The BChE isolated from blood plasma was digested with pepsin and analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS). The method was evaluated by using synthetic peptides and by comparison to the enzymatic activity Ellman's assay. The minimum concentration of VR exposure, resulting in detectable VR-adduct, was 0.2 ng/mL, which corresponded to the relative BChE inhibition of less than 2%. Adduct formation assessment was performed via monitoring of decrease in non-modified peptide LC-MS/MS signal and increase in VR-modified peptide signal. The designed approach was tested in a pilot study with 5 blood samples from healthy volunteers. Mass spectrometry-based method for BChE adduct formation was found to be in agreement with Ellman's inhibition assay, so the method is applicable for direct BChE inhibition assessment.

AB - Organophosphates’ toxic effect causes covalent binding to serine-198 in the active site of human plasma butyrylcholinesterase (BChE) with loss of enzymatic function (covalent inhibition). Mass spectrometric detection of modified FGESAGAAS peptide at the active site is a powerful exposure biomarker tool. The aim of this study was to develop mass spectrometry-based method for BChE adduct formation screening, avoiding the use of standard peptides. Immunomagnetic separation of proteins from plasma was optimized. Commercially available anti-butyrylcholinesterase monoclonal antibodies, immobilized on magnetic beads, resulted in stable and reusable affinity sorbent. The method was tested on horse serum BChE and real human plasma from healthy donors, treated with Russian VX (VR). The BChE isolated from blood plasma was digested with pepsin and analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS). The method was evaluated by using synthetic peptides and by comparison to the enzymatic activity Ellman's assay. The minimum concentration of VR exposure, resulting in detectable VR-adduct, was 0.2 ng/mL, which corresponded to the relative BChE inhibition of less than 2%. Adduct formation assessment was performed via monitoring of decrease in non-modified peptide LC-MS/MS signal and increase in VR-modified peptide signal. The designed approach was tested in a pilot study with 5 blood samples from healthy volunteers. Mass spectrometry-based method for BChE adduct formation was found to be in agreement with Ellman's inhibition assay, so the method is applicable for direct BChE inhibition assessment.

KW - Adducts

KW - Cholinesterase

KW - Covalent inhibition

KW - Immunomagnetic separation

KW - Organophosphorus compound

KW - Target proteomics

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U2 - 10.1016/j.talanta.2019.01.059

DO - 10.1016/j.talanta.2019.01.059

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EP - 382

JO - Talanta

JF - Talanta

SN - 0039-9140

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