title: Chemical labeling of electrochemically cleaved peptides.
abstract: BACKGROUND
  Cleavage of peptide bonds C-terminal to tyrosine and tryptophan after electrochemical oxidation may become a complementary approach to chemical and enzymatic cleavage. A chemical labeling approach specifically targeting reactive cleavage products is presented here and constitutes a promising first step towards the development of a new proteomics workflow.
  
  
  METHODS
  Hexylamine was used to react with the spirolactone moieties generated after electrochemical oxidation and cleavage of tripeptides. The influence of pH and reaction time on the yield was determined and the excess of tagging reagent was optimized. Selective detection of the tagged cleavage products was achieved by precursor ion scanning in a triple quadrupole mass spectrometer.
  
  
  RESULTS
  Optimal labeling was reached under aqueous conditions when working at pH 10 with a reaction time of 0.5 min. The excess of hexylamine over spirolactone groups can be significantly decreased by working under non-aqueous conditions in pure acetonitrile to prevent spirolactone hydrolysis. The specific formation of hexylamine-containing y(1) reporter ions generated by collision-induced dissociation (CID) tandem mass spectrometry (MS/MS) allows for selective detection by precursor ion scanning of the cleaved and labeled peptides.
  
  
  CONCLUSIONS
  This work presents a method for selective labeling and detection of electrochemically cleaved Tyr- and Trp-containing peptides for which reaction conditions have been optimized with hexylamine as labeling agent. This workflow offers new possibilities for electrochemical oxidation, cleavage and labeling of peptides and proteins.
authors: Analytical Biochemistry and Mass Spectrometry Core Facility, Department of Pharmacy, University of Groningen, Groningen, The Netherlands.
authors: Julien
authors: J
authors: Roeser
authors: Niels F A
authors: NF
authors: Alting
authors: Hjalmar P
authors: HP
authors: Permentier
authors: Andries P
authors: AP
authors: Bruins
authors: Rainer P H
authors: RP
authors: Bischoff
mesh_terms: D000588:Amines / Q000737:chemistry; D055664:Electrochemical Techniques / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D009842:Oligopeptides / Q000032:analysis* / Q000737:chemistry*; D010084:Oxidation-Reduction; D013148:Spironolactone / Q000737:chemistry; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014364:Tryptophan / Q000737:chemistry; D014443:Tyrosine / Q000737:chemistry
chemical_list: D000588:Amines; D009842:Oligopeptides; D013148:Spironolactone; D014443:Tyrosine; D014364:Tryptophan; C007940:hexylamine