title: Proteomic analysis of cellular soluble proteins from human bronchial smooth muscle cells by combining nondenaturing micro 2DE and quantitative LC-MS/MS. 2. Similarity search between protein maps for the analysis of protein complexes.
abstract: Human bronchial smooth muscle cell soluble proteins were analyzed by a combined method of nondenaturing micro 2DE, grid gel-cutting, and quantitative LC-MS/MS and a native protein map was prepared for each of the identified 4323 proteins [1]. A method to evaluate the degree of similarity between the protein maps was developed since we expected the proteins comprising a protein complex would be separated together under nondenaturing conditions. The following procedure was employed using Excel macros; (i) maps that have three or more squares with protein quantity data were selected (2328 maps), (ii) within each map, the quantity values of the squares were normalized setting the highest value to be 1.0, (iii) in comparing a map with another map, the smaller normalized quantity in two corresponding squares was taken and summed throughout the map to give an "overlap score," (iv) each map was compared against all the 2328 maps and the largest overlap score, obtained when a map was compared with itself, was set to be 1.0 thus providing 2328 "overlap factors," (v) step (iv) was repeated for all maps providing 2328 × 2328 matrix of overlap factors. From the matrix, protein pairs that showed overlap factors above 0.65 from both protein sides were selected (431 protein pairs). Each protein pair was searched in a database (UniProtKB) on complex formation and 301 protein pairs, which comprise 35 protein complexes, were found to be documented. These results demonstrated that native protein maps and their similarity search would enable simultaneous analysis of multiple protein complexes in cells.
authors.affiliation: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, P. R. China.|Guangdong Provincial Key Laboratory of Fermentation and Enzyme Engineering, South China University of Technology, Guangzhou, P. R. China.|Key Laboratory of Industrial Biotechnology of Guangdong Higher Education Institutes, South China University of Technology, Guangzhou, P. R. China.
authors.forename: Ya
authors.initials: Y
authors.lastname: Jin
authors.affiliation: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, P. R. China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Yuan
authors.affiliation: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, P. R. China.
authors.forename: Jun
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Faculty of Science, Ehime University, Matsuyama, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Manabe
authors.affiliation: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, P. R. China.|Key Laboratory of Industrial Biotechnology of Guangdong Higher Education Institutes, South China University of Technology, Guangzhou, P. R. China.
authors.forename: Wen
authors.initials: W
authors.lastname: Tan
mesh_terms: D002460:Cell Line; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D006801:Humans; D046912:Multiprotein Complexes / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000737:chemistry; D011506:Proteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D046912:Multiprotein Complexes; D011506:Proteins
keywords: Cellular proteins; Native protein map; Nondenaturing micro 2DE; Protein complex; Quantitative LC-MS/MS