title: Modification of human serum albumin by the nerve agent VX: microbore liquid chromatography/electrospray ionization high-resolution time-of-flight tandem mass spectrometry method for detection of phosphonylated tyrosine and novel cysteine containing disulfide adducts.
abstract: BACKGROUND
  Organophosphorus nerve agents still constitute a considerable threat to the health of military personnel and the civilian population. Long-term biomarkers are crucial for reliable verification of exposure to banned substances. Therefore, current research focuses on identification of endogenous protein targets showing covalent modifications by organophosphorus nerve agents (adducts).
  
  
  METHODS
  Purified human serum albumin and human plasma were incubated with the nerve agent VX followed by enzymatic proteolysis with pronase. Resulting peptide cleavage products were separated by microbore liquid chromatography (μLC) online coupled to positive electrospray ionization (ESI) with subsequent high-resolution time-of-flight tandem mass spectrometry (HR MS/MS) allowing identification of known and novel adducts.
  
  
  RESULTS
  In addition to known phosphonylation of various tyrosine residues, albumin was found to be modified at diverse cysteine residues by covalent attachment of the leaving group of VX. These novel disulfide adducts were cleaved from at least two regions of the intact protein as dipeptides containing cysteine and proline either as CP or PC. A rapid and sensitive method was developed for simultaneous detection of the diverse covalent modifications of human albumin by VX.
  
  
  CONCLUSIONS
  Identification of the novel leaving group adducts with human albumin expands the basic knowledge on molecular toxicology of the nerve agent VX. Furthermore, the presented μLC/ESI HR MS/MS method might be of relevance for verification of VX poisoning. Copyright © 2016 John Wiley & Sons, Ltd.
authors.affiliation: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, Munich, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Kranawetvogl
authors.affiliation: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, Munich, Germany.
authors.forename: Franz
authors.initials: F
authors.lastname: Worek
authors.affiliation: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, Munich, Germany.
authors.forename: Horst
authors.initials: H
authors.lastname: Thiermann
authors.affiliation: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, Munich, Germany. haraldjohn@bundeswehr.org.
authors.forename: Harald
authors.initials: H
authors.lastname: John
mesh_terms: D003545:Cysteine / Q000737:chemistry*; D004220:Disulfides / Q000737:chemistry; D006801:Humans; D000067397:Nerve Agents / Q000737:chemistry*; D009946:Organothiophosphorus Compounds / Q000737:chemistry*; D010766:Phosphorylation; D000075462:Serum Albumin, Human / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014443:Tyrosine / Q000737:chemistry*
chemical_list: D003545:Cysteine; D004220:Disulfides; D000067397:Nerve Agents; D009946:Organothiophosphorus Compounds; D000075462:Serum Albumin, Human; D014443:Tyrosine; C009680:VX-agent