title: Improving basic and membrane protein MS detection of the culture filtrate proteins from Mycobacterium tuberculosis H37Rv by biomimetic affinity prefractionation.
abstract: The culture filtrate proteins (CFPs) from Mycobacterium tuberculosis have been shown to induce protective immune responses in human and animal models, making them a promising source of candidate targets for tuberculosis drugs, vaccines, and diagnostics. The constituents of the M. tuberculosis CFP proteome are complex and vary with growth conditions. To effectively profile CFPs, gel-based prefractionation is usually performed before MS analysis. In this study, we describe a novel prefractionation approach by which the proteome is divided into seven partially overlapping fractions by biomimetic affinity chromatography (BiAC) using a six-column cascade. The LC-MS/MS analysis of individual fractions identified a total of 541 CFPs, including 61 first-time identifications. Notably, ∼1/3 (20/61) of these novel CFPs are membrane proteins, among which nine proteins have 2-14 transmembrane domains. In addition, ∼1/4 (14/61) of the CFPs are basic proteins with pI values greater than 9.0. Our data demonstrate that biomimetic affinity chromatography prefractionation markedly improves protein detection by LC-MS/MS, and the coverage of basic and hydrophobic proteins in particular is remarkably increased.
authors.affiliation: State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Guorong
authors.initials: G
authors.lastname: Ma
authors.affiliation: State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Kang
authors.initials: K
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Fuqiang
authors.initials: F
authors.lastname: Zhou
authors.affiliation: Clinical Laboratory of the Second Affiliated Hospital, Lanzhou University, Lanzhou, P. R. China.
authors.forename: Lina
authors.initials: L
authors.lastname: Gao
authors.affiliation: Department of Immunology and Microbiology, School of Medicine, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Zhanqiang
authors.initials: Z
authors.lastname: Sun
authors.affiliation: Shanghai HyCharm Inc., Shanghai, P. R. China.
authors.forename: Li
authors.initials: L
authors.lastname: Deng
authors.affiliation: Department of Immunology and Microbiology, School of Medicine, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Shulin
authors.initials: S
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.|Engineering Research Center of Cell and Therapeutic Antibody, Ministry of Education, Shanghai, P. R. China.
authors.forename: Rongxiu
authors.initials: R
authors.lastname: Li
keywords: Biomimetic affinity chromatography; Culture filtrate proteins; LC-MS/MS; Mycobacterium tuberculosis