期刊
METHODS
卷 56, 期 2, 页码 161-165出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymeth.2011.10.013
关键词
Receptor-ligand; Chemical crosslinking; Immunoaffinity purification; Differential mass spectrometry
资金
- WCU of National Research Foundation (NRF) of Korea [R31-2008-000-10103-0]
- government (MEST)
- 21C Frontier Functional Proteomics Program [FPR-10A1-034]
- Kangwon National University
- Seoul RBD Program [10035353]
Protein receptor-ligand interactions play important roles in mediating enzyme catalysis, signal transduction, and other protein functions. Immunoaffinity purification followed by mass spectrometry analysis is a common method for identifying protein receptor-ligand complexes. However, it is difficult to distinguish between specific protein binding partners and non-specifically bound proteins that co-purify with the complex. In addition, weakly interacting binding partners may dissociate from the protein receptor-ligand complexes during immunoaffinity purification. The combination of chemical crosslinking, affinity purification, and differential mass spectrometry analysis provides a direct method for capturing stable, weak, and transient protein interactions that occur in vivo and in vitro. This approach enables the identification of functional receptor-ligand binding partners with high confidence. Herein, we describe a differential mass spectrometry approach coupled with in situ chemical crosslinking and immunoaffinity purification for identifying receptor-ligand binding partners. In particular, we identified a functional, counter-ligand structure of the natural killer cell p30-related protein. (C) 2011 Elsevier Inc. All rights reserved.
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