PMID: 6413514Oct 1, 1983Paper

Detection of vinculin-binding proteins with an 125I-vinculin gel overlay technique

The Journal of Cell Biology
J J Otto

Abstract

Vinculin is an adhesion plaque component localized on the cytoplasmic side of the cell membrane where stress fibers end. To detect vinculin-binding proteins, we have developed an 125I-vinculin gel overlay method. SDS PAGE was used to separate different protein preparations. After fixing the proteins in the gel with methanol-acetic acid, the SDS was removed with ethanol and the proteins renatured in buffer. The gel was then incubated with 125I-vinculin. After extensive washing to remove nonspecifically associated label, the gel was dried and autoradiographed. Chick embryo fibroblasts, their Rous sarcoma virus transformants, and HeLa cells were found to contain two proteins (Mr 220,000 and 130,000) that bound 125I-vinculin strongly and another (Mr 42,000) that bound it moderately. The 130,000-mol-wt protein was identified as vinculin itself, which suggests that it may self-associate. The 42,000-mol-wt protein was identified as actin with which vinculin is known to interact. The identity of the 220,000-mol-wt protein is not known. It is not cellular fibronectin, myosin, or filamin. When fibroblast proteins were separated into Triton X-100 soluble and insoluble fractions, most of the vinculin and the 220,000-mol-wt protein was foun...Continue Reading

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Citations

Jan 1, 1990·Cell Motility and the Cytoskeleton·M E Groesch, J J Otto
Jan 1, 1990·Cell Motility and the Cytoskeleton·J J Otto
Jan 1, 1990·Cell Motility and the Cytoskeleton·M C Beckerle, R K Yeh
Jan 1, 1991·Cell Motility and the Cytoskeleton·C P ChiaE J Luna
Jan 1, 1995·Cell Motility and the Cytoskeleton·K KatohK Fujiwara
Dec 2, 1990·Cell Differentiation and Development : the Official Journal of the International Society of Developmental Biologists·K BurridgeC Turner
Apr 16, 2002·Cellular Microbiology·Toshio KodamaTakeshi Honda
Oct 15, 1991·Proceedings of the National Academy of Sciences of the United States of America·K HerrenknechtR Kemler
Apr 15, 1993·Proceedings of the National Academy of Sciences of the United States of America·J Hagmann
Mar 1, 1985·The Journal of Cell Biology·G C RosenfeldJ Bryan
Oct 1, 1985·The Journal of Cell Biology·B GeigerT Volberg
Nov 1, 1986·The Journal of Cell Biology·M C Beckerle
Mar 1, 1987·The Journal of Cell Biology·J D Siliciano, S W Craig
Aug 1, 1988·The Journal of Cell Biology·A M BelkinV E Koteliansky
Aug 1, 1989·The Journal of Cell Biology·J D ZieskeI K Gipson
Dec 1, 1989·The Journal of Cell Biology·P JonesD R Critchley
May 1, 1990·The Journal of Cell Biology·G H NuckollsK Burridge
Aug 1, 1990·The Journal of Cell Biology·C A OteyK Burridge
Sep 1, 1990·The Journal of Cell Biology·C E TurnerK Burridge
Aug 1, 1991·The Journal of Cell Biology·R J Barstead, R H Waterston
Mar 1, 1992·The Journal of Cell Biology·A W CrawfordM C Beckerle
Jul 1, 1984·The Journal of Cell Biology·P Mangeat, K Burridge
May 31, 2007·Genes to Cells : Devoted to Molecular & Cellular Mechanisms·Kazuo KatohShigeo Ookawara
Feb 1, 1986·Experimental Cell Research·G R MatyasD J Morré

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