Mass spectral characterization of organophosphate-labeled, tyrosine-containing peptides

Characteristic mass fragments and a new binding motif for organophosphates

Lawrence M Schopfer, Hasmik Grigoryan, Bin Li, Florian Nachon, Patrick Masson, Oksana Lockridge

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

We have identified organophosphorus agent (OP)-tyrosine adducts on 12 different proteins labeled with six different OP. Labeling was achieved by treating pure proteins with up to 40-fold molar excess of OP at pH 8-8.6. OP-treated proteins were digested with trypsin, and peptides were separated by HPLC. Fragmentation patterns for 100 OP-peptides labeled on tyrosine were determined in the mass spectrometer. The goals of the presentworkwere (1) to determine thecommonfeatures of the OP-reactive tyrosines, and (2) to describe non-sequence MSMS fragments characteristic of OP-tyrosine peptides. Characteristic ions at 272 and 244amu for tyrosine-OP immonium ionswere nearly always present in the MSMS spectrum of peptides labeled on tyrosine by chlorpyrifos-oxon. Characteristic fragments also appeared fromthe parent ions that had been labeled with diisopropylfluorophosphate (216 amu), sarin (214 amu), soman (214 amu) or FP-biotin (227, 312, 329, 691 and 708 amu). In contrast to OP-reactive serines, which lie in the consensus sequence GXSXG, the OP-reactive tyrosines have no consensus sequence. Their common feature is the presence of nearby positively charged residues that activate the phenolic hydroxyl group. The significance of these findings is the recognition of a new binding motif for OP to proteins that have no active site serine. Modified peptides are difficult to findwhentheOPbears no radiolabel and no tag. The characteristicMSMS fragment ions are valuable because they are identifiers for OP-tyrosine, independent of the peptide.

Original languageEnglish (US)
Pages (from-to)1297-1311
Number of pages15
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume878
Issue number17-18
DOIs
StatePublished - May 15 2010

Fingerprint

Organophosphates
Tyrosine
Peptides
Consensus Sequence
Ions
Serine
Proteins
Sarin
Soman
Isoflurophate
Hydroxyl Radical
Trypsin
Catalytic Domain
High Pressure Liquid Chromatography
Mass spectrometers
Labeling

Keywords

  • Characteristic ions
  • Covalent bond
  • Mass spectrometry
  • Organophosphorus
  • Tyrosine

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry
  • Cell Biology

Cite this

@article{4ae6de5bdc524e77bb34c7784b760691,
title = "Mass spectral characterization of organophosphate-labeled, tyrosine-containing peptides: Characteristic mass fragments and a new binding motif for organophosphates",
abstract = "We have identified organophosphorus agent (OP)-tyrosine adducts on 12 different proteins labeled with six different OP. Labeling was achieved by treating pure proteins with up to 40-fold molar excess of OP at pH 8-8.6. OP-treated proteins were digested with trypsin, and peptides were separated by HPLC. Fragmentation patterns for 100 OP-peptides labeled on tyrosine were determined in the mass spectrometer. The goals of the presentworkwere (1) to determine thecommonfeatures of the OP-reactive tyrosines, and (2) to describe non-sequence MSMS fragments characteristic of OP-tyrosine peptides. Characteristic ions at 272 and 244amu for tyrosine-OP immonium ionswere nearly always present in the MSMS spectrum of peptides labeled on tyrosine by chlorpyrifos-oxon. Characteristic fragments also appeared fromthe parent ions that had been labeled with diisopropylfluorophosphate (216 amu), sarin (214 amu), soman (214 amu) or FP-biotin (227, 312, 329, 691 and 708 amu). In contrast to OP-reactive serines, which lie in the consensus sequence GXSXG, the OP-reactive tyrosines have no consensus sequence. Their common feature is the presence of nearby positively charged residues that activate the phenolic hydroxyl group. The significance of these findings is the recognition of a new binding motif for OP to proteins that have no active site serine. Modified peptides are difficult to findwhentheOPbears no radiolabel and no tag. The characteristicMSMS fragment ions are valuable because they are identifiers for OP-tyrosine, independent of the peptide.",
keywords = "Characteristic ions, Covalent bond, Mass spectrometry, Organophosphorus, Tyrosine",
author = "Schopfer, {Lawrence M} and Hasmik Grigoryan and Bin Li and Florian Nachon and Patrick Masson and Oksana Lockridge",
year = "2010",
month = "5",
day = "15",
doi = "10.1016/j.jchromb.2009.07.026",
language = "English (US)",
volume = "878",
pages = "1297--1311",
journal = "Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences",
issn = "1570-0232",
publisher = "Elsevier",
number = "17-18",

}

TY - JOUR

T1 - Mass spectral characterization of organophosphate-labeled, tyrosine-containing peptides

T2 - Characteristic mass fragments and a new binding motif for organophosphates

AU - Schopfer, Lawrence M

AU - Grigoryan, Hasmik

AU - Li, Bin

AU - Nachon, Florian

AU - Masson, Patrick

AU - Lockridge, Oksana

PY - 2010/5/15

Y1 - 2010/5/15

N2 - We have identified organophosphorus agent (OP)-tyrosine adducts on 12 different proteins labeled with six different OP. Labeling was achieved by treating pure proteins with up to 40-fold molar excess of OP at pH 8-8.6. OP-treated proteins were digested with trypsin, and peptides were separated by HPLC. Fragmentation patterns for 100 OP-peptides labeled on tyrosine were determined in the mass spectrometer. The goals of the presentworkwere (1) to determine thecommonfeatures of the OP-reactive tyrosines, and (2) to describe non-sequence MSMS fragments characteristic of OP-tyrosine peptides. Characteristic ions at 272 and 244amu for tyrosine-OP immonium ionswere nearly always present in the MSMS spectrum of peptides labeled on tyrosine by chlorpyrifos-oxon. Characteristic fragments also appeared fromthe parent ions that had been labeled with diisopropylfluorophosphate (216 amu), sarin (214 amu), soman (214 amu) or FP-biotin (227, 312, 329, 691 and 708 amu). In contrast to OP-reactive serines, which lie in the consensus sequence GXSXG, the OP-reactive tyrosines have no consensus sequence. Their common feature is the presence of nearby positively charged residues that activate the phenolic hydroxyl group. The significance of these findings is the recognition of a new binding motif for OP to proteins that have no active site serine. Modified peptides are difficult to findwhentheOPbears no radiolabel and no tag. The characteristicMSMS fragment ions are valuable because they are identifiers for OP-tyrosine, independent of the peptide.

AB - We have identified organophosphorus agent (OP)-tyrosine adducts on 12 different proteins labeled with six different OP. Labeling was achieved by treating pure proteins with up to 40-fold molar excess of OP at pH 8-8.6. OP-treated proteins were digested with trypsin, and peptides were separated by HPLC. Fragmentation patterns for 100 OP-peptides labeled on tyrosine were determined in the mass spectrometer. The goals of the presentworkwere (1) to determine thecommonfeatures of the OP-reactive tyrosines, and (2) to describe non-sequence MSMS fragments characteristic of OP-tyrosine peptides. Characteristic ions at 272 and 244amu for tyrosine-OP immonium ionswere nearly always present in the MSMS spectrum of peptides labeled on tyrosine by chlorpyrifos-oxon. Characteristic fragments also appeared fromthe parent ions that had been labeled with diisopropylfluorophosphate (216 amu), sarin (214 amu), soman (214 amu) or FP-biotin (227, 312, 329, 691 and 708 amu). In contrast to OP-reactive serines, which lie in the consensus sequence GXSXG, the OP-reactive tyrosines have no consensus sequence. Their common feature is the presence of nearby positively charged residues that activate the phenolic hydroxyl group. The significance of these findings is the recognition of a new binding motif for OP to proteins that have no active site serine. Modified peptides are difficult to findwhentheOPbears no radiolabel and no tag. The characteristicMSMS fragment ions are valuable because they are identifiers for OP-tyrosine, independent of the peptide.

KW - Characteristic ions

KW - Covalent bond

KW - Mass spectrometry

KW - Organophosphorus

KW - Tyrosine

UR - http://www.scopus.com/inward/record.url?scp=77954087882&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=77954087882&partnerID=8YFLogxK

U2 - 10.1016/j.jchromb.2009.07.026

DO - 10.1016/j.jchromb.2009.07.026

M3 - Article

VL - 878

SP - 1297

EP - 1311

JO - Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences

JF - Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences

SN - 1570-0232

IS - 17-18

ER -