Premium
Utility of electrophoretically derived protein mass estimates as additional constraints in proteome analysis of human serum based on MS/MS analysis
Author(s) -
Kim Jin Young,
Lee Jeong Hwa,
Park Gun Wook,
Cho Kun,
Kwon KyungHoon,
Park Young Mok,
Cho Sang Yun,
Paik YoungKi,
Yoo Jong Shin
Publication year - 2005
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200401220
Subject(s) - proteome , chromatography , chemistry , mass spectrometry , human proteome project , proteomics , trypsin , blood proteins , peptide , human plasma , computational biology , biology , biochemistry , gene , enzyme
Abstract The proteome of a HUPO human serum reference sample was analyzed using multidimensional separation techniques at both the protein and the peptide levels. To eliminate false‐positive identifications from the search results, we employed a data filtering method using molecular weight (MW) correlations derived from denaturing 1‐DE. First, the six most abundant serum proteins were removed from the sample using immunoaffinity chromatography. 1‐DE was then used to fractionate the remaining serum proteins according to the MW. Gel bands were isolated and in‐gel digested with trypsin, and the resulting peptides were analyzed by 2‐D LC/ESI‐MS/MS. A SEQUEST search using the MS/MS results identified 494 proteins. Of these, 202 were excluded formally using protein data filtering as they were single‐assignment proteins and their theoretical and electrophoretically‐derived MWs did not correlate at high confidence. To evaluate this method, the results were compared with those of 1‐D LC/MALDI‐TOF/TOF and HUPO Plasma Proteome Project analyses. Our data filtering approach proved valuable in analysis of complex, large‐scale proteomes such as human serum.