期刊
JOURNAL OF MOLECULAR BIOLOGY
卷 418, 期 3-4, 页码 134-144出版社
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.jmb.2011.12.053
关键词
ubiquitin; linkage; phage display; immunoglobulin; X-ray crystallography
资金
- National Institutes of Health, National Institute of General Medical Sciences
- Howard Hughes Medical Institute
- Office of Science, Office of Basic Energy Sciences, of U.S. Department of Energy [DE-AC02-05CH11231]
Polyubiquitination is an essential posttranslational modification that plays critical roles in cellular signaling. PolyUb (polyubiquitin) chains are formed by linking the carboxyl-terminus of one Ub (ubiquitin) subunit to either a lysine residue or the amino-terminus of an adjacent Ub. Linkage through the amino-terminus results in linear polyubiquitination that has recently been demonstrated to be a key step in nuclear factor kappa B activation; however, tools to study linear chains have been lacking. We therefore engineered a linear-linkage-specific antibody that is functional in Western blot, immunoprecipitation, and immunofluorescence applications. A crystal structure of the linear-linkage-specific antibody Fab fragment in complex with linear diubiquitin provides molecular insight into the nature of linear chain specificity. We use the antibody to demonstrate that linear polyUb is up-regulated upon tumor necrosis factor a stimulation of cells, consistent with a critical role in nuclear factor kappa B signaling. This antibody provides an essential tool for further investigation of the function of linear chains. (C) 2011 Published by Elsevier Ltd.
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