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BioMed Research International
Volume 2015, Article ID 134050, 21 pages
http://dx.doi.org/10.1155/2015/134050
Research Article

Identification of Glioblastoma Phosphotyrosine-Containing Proteins with Two-Dimensional Western Blotting and Tandem Mass Spectrometry

Tianyao Guo,1,2,3 Xiaowei Wang,1,2,3 Maoyu Li,1,2,3 Haiyan Yang,1,2,3 Ling Li,1 Fang Peng,1,2,3 and Xianquan Zhan1,2,3,4

1Key Laboratory of Cancer Proteomics of Chinese Ministry of Health, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
2Hunan Engineering Laboratory for Structural Biology and Drug Design, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
3State Local Joint Engineering Laboratory for Anticancer Drugs, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan 410008, China
4The State Key Laboratory of Medical Genetics, Central South University, 88 Xiangya Road, Changsha, Hunan 410008, China

Received 11 June 2014; Revised 25 August 2014; Accepted 25 August 2014

Academic Editor: Fa-Yun Che

Copyright © 2015 Tianyao Guo et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

To investigate the presence of, and the potential biological roles of, protein tyrosine phosphorylation in the glioblastoma pathogenesis, two-dimensional gel electrophoresis- (2DGE-) based Western blotting coupled with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) analysis was used to detect and identify the phosphotyrosine immunoreaction-positive proteins in a glioblastoma tissue. MS/MS and Mascot analyses were used to determine the phosphotyrosine sites of each phosphopeptide. Protein domain and motif analysis and systems pathway analysis were used to determine the protein domains/motifs that contained phosphotyrosine residue and signal pathway networks to clarify the potential biological functions of protein tyrosine phosphorylation. A total of 24 phosphotyrosine-containing proteins were identified. Each phosphotyrosine-containing protein contained at least one tyrosine kinase phosphorylation motif and a certain structural and functional domains. Those phosphotyrosine-containing proteins were involved in the multiple signal pathway systems such as oxidative stress, stress response, and cell migration. Those data show 2DGE-based Western blotting, MS/MS, and bioinformatics are a set of effective approaches to detect and identify glioblastoma tyrosine-phosphorylated proteome and to effectively rationalize the biological roles of tyrosine phosphorylation in the glioblastoma biological systems. It provides novel insights regarding tyrosine phosphorylation and its potential role in the molecular mechanism of a glioblastoma.