Acid-labile surfactant improves in-sodium dodecyl sulfate polyacrylamide gel protein digestion for matrix-assisted laser desorption/ionization mass spectrometric peptide mapping.
Mass spectrometry (MS) together with genome database searches serves as a powerful tool for the identification of proteins. In proteome analysis, mixtures of cellular proteins are usually separated by sodium dodecyl sulfate (SDS) polyacrylamide gel-based two-dimensional gel electrophoresis (2-DE) or one-dimensional gel electrophoresis (1-DE), and in-gel digested by a specific protease. In-gel protein digestion is one of the critical steps for sensitive protein identification by these procedures. Efficient protein digestion is required for obtaining peptide peaks necessary for protein identification by MS. This paper reports a remarkable improvement of protein digestion in SDS polyacrylamide gels using an acid-labile surfactant, sodium 3-[(2-methyl-2-undecyl-1,3-dioxolan-4-yl)methoxy]-1-propanesulfonate (ALS). Pretreatment of gel pieces containing protein spots separated by 2-DE with a small amount of ALS prior to trypsin digestion led to increases in the digested peptides eluted from the gels. Consistently, treatment of gel pieces containing silver-stained standard proteins and those separated from tissue extracts resulted in the detection of increased numbers of peptide peaks in spectra obtained by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOFMS). Hence the present protocol with ALS provides a useful strategy for sensitive protein identification by MS.
Nomura
Eiko
E
Division of Signal Transduction, Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma 630-0192, Japan.
Katsuta
Kazuhiro
K
Ueda
Tomoko
T
Toriyama
Michinori
M
Mori
Tatsuya
T
Inagaki
Naoyuki
N
D000476:Alkanesulfonates / Q000737:chemistry*; D000818:Animals; D002478:Cells, Cultured; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D010449:Peptide Mapping / Q000379:methods*; D040901:Proteomics / Q000379:methods; D051381:Rats; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D013501:Surface-Active Agents / Q000737:chemistry*
D000476:Alkanesulfonates; D013501:Surface-Active Agents; D012967:Sodium Dodecyl Sulfate; C028913:dodecyl sulfate