title: Combination of Edman degradation of peptides with liquid chromatography/mass spectrometry workflow for peptide identification in bottom-up proteomics.
abstract: BACKGROUND
  High-throughput methods of proteomics are essential for identification of proteins in a cell or tissue under certain conditions. Most of these methods require tandem mass spectrometry (MS/MS). A multidimensional approach including predictive chromatography and partial chemical degradation could be a valuable alternative and/or addition to MS/MS.
  
  
  METHODS
  In the proposed strategy peptides are identified in a three-dimensional (3D) search space consisting of retention time (RT), mass, and reduced mass after one-step partial Edman degradation. The strategy was evaluated in silico for two databases: baker's yeast and human proteins. Rates of unambiguous identifications were estimated for mass accuracies from 0.001 to 0.05 Da and RT prediction accuracies from 0.1 to 5 min. Rates of Edman reactions were measured for test peptides.
  
  
  RESULTS
  A 3D description of proteolytic peptides allowing unambiguous identification without employing MS/MS of up to 95% and 80% of tryptic peptides from the yeast and human proteomes, respectively, was considered. Further extension of the search space to a four-dimensional one by incorporating the second N-terminal amino acid residue as the fourth dimension was also considered and was shown to result in up to 90% of human peptides being identified unambiguously.
  
  
  CONCLUSIONS
  The proposed 3D search space can be a useful alternative to MS/MS-based peptide identification approach. Experimental implementations of the proposed method within the on-line liquid chromatography/mass spectrometry (LC/MS) and off-line matrix-assisted laser desorption/ionization (MALDI) workflows are in progress.
authors.affiliation: Institute for Energy Problems of Chemical Physics, Russian Academy of Sciences, Moscow, Russia.
authors.forename: Anna A
authors.initials: AA
authors.lastname: Lobas
authors.forename: Anatoly N
authors.initials: AN
authors.lastname: Verenchikov
authors.forename: Anton A
authors.initials: AA
authors.lastname: Goloborodko
authors.forename: Lev I
authors.initials: LI
authors.lastname: Levitsky
authors.forename: Mikhail V
authors.initials: MV
authors.lastname: Gorshkov
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003198:Computer Simulation; D030562:Databases, Protein; D006801:Humans; D009943:Organophosphorus Compounds / Q000737:chemistry*; D010446:Peptide Fragments / Q000032:analysis* / Q000737:chemistry; D011506:Proteins / Q000032:analysis / Q000737:chemistry; D040901:Proteomics / Q000379:methods*; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis / Q000737:chemistry; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: C043517:2-(4-isothiocyanatophenoxy)-1,3,2-dioxaphosphinene 2-oxide; D009943:Organophosphorus Compounds; D010446:Peptide Fragments; D011506:Proteins; D029701:Saccharomyces cerevisiae Proteins