Proteomics based on peptide fractionation by SDS-free PAGE

作者：Ramos Yassel; Gutierrez Elain; Machado Yoan; Sanchez Aniel; Castellanos Serra Lila; Gonzalez Luis J; Fernandez de Cossio Jorge; Perez Riverol Yasset; Betancourt Lazaro; Gil Jeovanis; Padron Gabriel; Besada Vladimir^*

来源：Journal of Proteome Research, 2008, 7(6): 2427-2434.

DOI：10.1021/pr700840y

• fractionation
• peptide
• mass spectrometry.
• proteomic
• protein
• electrophoresis
• SDS
• PAGE
• Neisseria meningitidis

Here we demonstrate the usefulness of peptide fractionation by SIDS-free polyacrylamide gel electrophoresis and its applicability to proteomics studies. In the absence of SIDS, the driving force for the electrophoretic migration toward the anode is supplied by negatively charged acidic aminoacid residues and other residues as phosphate, sulfate and sialic acid, while the resulting mobility depends on both the charge and the molecular mass of the peptides. A straightforward method was achieved for SDS-PAGE of proteins, enzyme digestion, peptide transfer and fractionation by SIDS-free PAGE, which was named dual-fractionation polyacrylamide gel electrophoresis (DF-PAGE). This method increases the number of identified proteins 2.5-fold with respect to the proteins identified after direct analysis, and more than 80% of assigned peptides were found in unique SDS-free gel slices. A vast majority of identified peptides (93%) have p/values below 7.0, and 7% have p/values between 7.0 and 7.35. Peptide digests that were derived from complex protein mixtures were in consequence simplified as peptides that are positively charged are not-recovered in the present conditions. The analysis of a, membrane protein extract from Neisseria meningitidis by this approach allowed the identification of 97 proteins, including low-abundance components.

• 出版日期2008-6