Phosphoric acid as a matrix additive for MALDI MS analysis of phosphopeptides and phosphoproteins.
Phosphopeptides are often detected with low efficiency by MALDI MS analysis of peptide mixtures. In an effort to improve the phosphopeptide ion response in MALDI MS, we investigated the effects of adding low concentrations of organic and inorganic acids during peptide sample preparation in 2,5-dihydroxybenzoic acid (2,5-DHB) matrix. Phosphoric acid in combination with 2,5-DHB matrix significantly enhanced phosphopeptide ion signals in MALDI mass spectra of crude peptide mixtures derived from the phosphorylated proteins alpha-casein and beta-casein. The beneficial effects of adding up to 1% phosphoric acid to 2,5-DHB were also observed in LC-MALDI-MS analysis of tryptic phosphopeptides of B. subtilis PrkC phosphoprotein. Finally, the mass resolution of MALDI mass spectra of intact proteins was significantly improved by using phosphoric acid in 2,5-DHB matrix.
Kjellström
Sven
S
Protein Research Group, Department of Biochemistry & Molecular Biology, University of Southern Denmark, DK-5230 Odense M, Denmark.
Jensen
Ole Nørregaard
ON
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry; D002364:Caseins / Q000737:chemistry; D005841:Gentisates / Q000737:chemistry; D006801:Humans; D007477:Ions / Q000737:chemistry; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis; D010449:Peptide Mapping / Q000379:methods*; D010748:Phosphopeptides / Q000032:analysis*; D010750:Phosphoproteins / Q000737:chemistry*; D010756:Phosphoric Acids / Q000737:chemistry*; D010766:Phosphorylation; D012680:Sensitivity and Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D014357:Trypsin / Q000737:chemistry
D001426:Bacterial Proteins; D002364:Caseins; D005841:Gentisates; D007477:Ions; D010446:Peptide Fragments; D010748:Phosphopeptides; D010750:Phosphoproteins; D010756:Phosphoric Acids; C030242:phosphoric acid; D014357:Trypsin; C010925:2,5-dihydroxybenzoic acid