Determination of protein adducts of organophosphorus nerve agents in blood plasma

Koryagina, N. L.; Savel’eva, E. I.; Karakashev, G. V.; Babakov, V. N.; Dubrovskii, Yaroslav A.; Ukolova, E. S.; Khlebnikova, N. S.; Murashko, E. A.; Koneva, V. Yu.; Ukolov, A. I.; Копейкин, В. А.; Radilov, A. S.

doi:10.1134/s1061934816080086

Cited by 14 publications

(5 citation statements)

References 17 publications

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“…Monoclonal HAH 002-01, also called 3E8, from BioPorto Diagnostics or from Thermo Fisher has been used by the Centers for Control and Prevention (Atlanta, GA) and the Federal Medical-Biological Agency of Russia (St. Petersburg) to immunopurify HuBChE from plasma. 1,16 Monoclonal 3E8 has been used in a magnetic electrochemical sensing platform to monitor exposure to organophosphorus agents. 17 Monoclonals mAb2 and B2 12-1 can also be used to immunopurify HuBChE from plasma, 8 although they are not commercially available.…”

Section: ■ Discussionmentioning

confidence: 99%

“…Other monoclonal antibodies to HuBChE can be used in place of B2 18-5 to capture HuBChE from plasma. Monoclonal HAH 002-01, also called 3E8, from BioPorto Diagnostics or from Thermo Fisher has been used by the Centers for Control and Prevention (Atlanta, GA) and the Federal Medical-Biological Agency of Russia (St. Petersburg) to immunopurify HuBChE from plasma. , Monoclonal 3E8 has been used in a magnetic electrochemical sensing platform to monitor exposure to organophosphorus agents …”

Section: Discussionmentioning

confidence: 99%

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Monoclonal Antibodies That Recognize Various Folding States of Pure Human Butyrylcholinesterase Can Immunopurify Butyrylcholinesterase from Human Plasma Stored at Elevated Temperatures

et al. 2016

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Human plasma to be analyzed for exposure to cholinesterase inhibitors is stored at 4 °C or lower to prevent denaturation of human butyrylcholinesterase (HuBChE), the biomarker of exposure. Currently published protocols immunopurify HuBChE using antibodies that bind native HuBChE before analysis by mass spectrometry. It is anticipated that the plasma collected from human casualties may be stored nonideally at elevated temperatures of up to 45 °C for days or maybe weeks. At 45 °C, the plasma loses 50% of its HuBChE activity in 8 days and 95% in 40 days. Our goal was to identify a set of monoclonal antibodies that could be used to immunopurify HuBChE from plasma stored at 45 °C. The folding states of pure human HuBChE stored at 4 and 45 °C and boiled at 100 °C were visualized on nondenaturing gels stained with Coomassie blue. Fully active pure HuBChE tetramers had a single band, but pure HuBChE stored at 45 °C had four bands, representing native, partly unfolded, aggregated, and completely denatured, boiled tetramers. The previously described monoclonal B2 18-5 captured native, partly unfolded, and aggregated HuBChE tetramers, whereas a new monoclonal, C191 developed in our laboratory, was found to selectively capture completely denatured, boiled HuBChE. The highest quantity of HuBChE protein was extracted from 45 °C heat-denatured human plasma when HuBChE was immunopurified with a combination of monoclonals B2 18-5 and C191. Using a mixture of these two antibodies in future emergency response assays may increase the capability to confirm exposure to cholinesterase inhibitors.

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Section: ■ Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Monoclonal Antibodies That Recognize Various Folding States of Pure Human Butyrylcholinesterase Can Immunopurify Butyrylcholinesterase from Human Plasma Stored at Elevated Temperatures

et al. 2016

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“…Unusually high quantities of pepsin are required to produce the nine-residue peptide, as pointed out by Fidder et al 14 and confirmed by others. 2 − 4 , 6 , 15 − 17 Digestion with lower quantities of pepsin yields a mixture of active site peptides, containing 12 residues or more. The 9-residue peptide is ideal for mass spectrometry analysis because the short peptide ionizes readily and yields a fragmentation spectrum that contains at least three characteristic fragment ions.…”

Section: Materials and Methodsmentioning

confidence: 99%

Use of Hupresin To Capture Red Blood Cell Acetylcholinesterase for Detection of Soman Exposure

et al. 2017

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Toxicity from acute exposure to nerve agents and organophosphorus toxicants is due to irreversible inhibition of acetylcholinesterase (AChE) in the nervous system. AChE in red blood cells is a surrogate for AChE in the nervous system. Previously we developed an immunopurification method to enrich red blood cell AChE (RBC AChE) as a biomarker of exposure. The goal of the present work was to provide an alternative RBC AChE enrichment strategy, by binding RBC AChE to Hupresin affinity gel. AChE was solubilized from frozen RBC by addition of 1% Triton X-100. Insoluble debris was removed by centrifugation. The red, but not viscous, RBC AChE solution was loaded on a Hupresin affinity column. Hemoglobin and other proteins were washed off with 3 M NaCl, while retaining AChE bound to Hupresin. Denatured AChE was eluted with 1% trifluoroacetic acid. The same protocol was used for 20 mL of RBC AChE inhibited with a soman model compound. The acid denatured protein was digested with pepsin and analyzed by liquid chromatography tandem mass spectrometry on a 6600 Triple-TOF mass spectrometer. A targeted method identified the aged soman adduct on serine 203 in peptide FGESAGAAS. It was concluded that Hupresin can be used to enrich soman-inhibited AChE solubilized from 8 mL of frozen human erythrocytes, yielding a quantity sufficient for detecting soman exposure.

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“…Previous studies by conventional LC-MS analysis have found that OPs can covalently bind to proteins such as butyryl-cholinesterase (BChE) [11][12][13], albumin [14][15][16], ceruloplasmin [17], and AChE. Recently, Pei and the co-workers identified several characteristic biomarkers of G&V-type chemical agents from albumin in vitro and in vivo by using Q-Orbitrap-MS analysis [18,19].…”

Section: Introductionmentioning

confidence: 99%

“…Moreover, as these OP‐protein adducts or metabolites have the characteristics of toxic agents and show a certain dose‐effect relationship, they are often used for retrospective verification and/or dose assessment of toxic agents. Previous studies by conventional LC‐MS analysis have found that OPs can covalently bind to proteins such as butyryl‐cholinesterase (BChE) [11–13], albumin [14–16], ceruloplasmin [17], and AChE. Recently, Pei and the co‐workers identified several characteristic biomarkers of G&V‐type chemical agents from albumin in vitro and in vivo by using Q‐Orbitrap‐MS analysis [18, 19].…”

Section: Introductionmentioning

confidence: 99%

Comprehensive insight on protein modification by V‐type agent: A chemical proteomic approach employing bioorthogonal reaction

Xing,

Wang,

Cui

et al. 2023

Proteomics

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Organophosphorus compounds (OPs) such as chemical agents and pesticides are posing critical threats to civilians due to their irreversible phosphonylation of diverse amino acids residues forming different protein adducts. However, traditional analytical approaches are quite limited in capturing the myriad of post‐translational events that affect protein functions, especially in identifying the low‐abundance OP adducts. Herein a systematic proteomic strategy based on a typical click‐enrich‐release‐identify bioorthogonal operation was firstly developed by employing an alkynyl‐tagged V‐type agent probe (AVP) and a biotin‐based azido‐enrichment linker (BTP‐N3). AVP targeting peptides from human serum albumin (HSA) or plasma were captured by BTP‐N3 via CuAAC click reaction, enriched by streptavidin beads, released by selective alkaline hydrolysis of phenacyl ester bond, and subsequently sequenced by LC‐MS/MS. This strategy has helped identifying 1115 unique OP adduction sites on 163 proteins in human plasma, and covers lots of OP adducts that cannot be achieved by traditional detection methods. The comprehensive coverage of novel OP substrates provided a general and sensitive approach to retrospective verification and/or dose assessment of toxic OPs.

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Determination of protein adducts of organophosphorus nerve agents in blood plasma

Cited by 14 publications

References 17 publications

Monoclonal Antibodies That Recognize Various Folding States of Pure Human Butyrylcholinesterase Can Immunopurify Butyrylcholinesterase from Human Plasma Stored at Elevated Temperatures

Monoclonal Antibodies That Recognize Various Folding States of Pure Human Butyrylcholinesterase Can Immunopurify Butyrylcholinesterase from Human Plasma Stored at Elevated Temperatures

Use of Hupresin To Capture Red Blood Cell Acetylcholinesterase for Detection of Soman Exposure

Comprehensive insight on protein modification by V‐type agent: A chemical proteomic approach employing bioorthogonal reaction

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