A new method for differential proteomic analysis utilizing native MS-electropherograms: a comparison between plasma and serum proteins by combining native PAGE, whole-gel slicing and quantitative LC-MS/MS

ABSTRACT: MS identification has long been used for PAGE-separated protein bands, but global and systematic quantitation utilizing MS after PAGE has remained rare and not been reported for native PAGE. Here we reported on a new method combining native PAGE, whole-gel slicing and quantitative LC-MS/MS aiming comparative analysis on not only protein abundance, but structures and interactions in different samples. A pair of human plasma and serum samples were used as test samples and separated on a native PAGE gel. Six lanes of each sample were cut, each lane was further sliced into thirty-five 1.1 mm x 1.1 mm squares and each square was subjected to standardized procedures of in-gel digestion and quantitative LC-MS/MS. The results, comprising 958 data rows that each contains the abundance information of a protein in one square in eleven gel lanes (one plasma lane excluded), showed satisfactory reproducibility of assignment and quantitation. A total of 315 proteins were assigned and for each protein, the abundance distributions in the plasma and serum gel lanes were reconstructed, respectively (named as ‘native MS-electropherograms’). Comparison of the electropherograms revealed significant plasma-vs-serum differences in 33 proteins (fold difference > 2 or < 0.5, p < 0.05), among which 18 showed differences in more than one squares. Many of the differences matched with accumulated knowledge on protein interactions and proteolysis involved in blood coagulation, complement and wound healing processes. We expect this method would be used to analyze changes or differences between proteomic samples, providing information on both protein abundance and structures.

INSTRUMENT(S): Synapt MS

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Plasma, Blood Serum

SUBMITTER: Ya Jin  

LAB HEAD: Ya Jin

PROVIDER: PXD006773 | Pride | 2017-09-08

REPOSITORIES: Pride