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Title: Two-dimensional gel electrophoresis of actin-binding proteins isolated by affinity chromatography from human skeletal muscle
Contributor(s): Coumans-Moens, Joelle (author)orcid ; Humphery-Smith, Ian (author); dos Remedios, Cristobal G (author)
Publication Date: 1997
DOI: 10.1002/elps.1150180709
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Abstract: In muscle cells actin exists as a mixture of monomeric (G-actin) and filamentous actin (F-actin) and ionic conditions strongly favor the formation of F-actin. The existence of unpolymerized actin depends, among other factors, on proteins that bind to G-actin, the so-called G-actin-binding proteins (G-ABPs). We have coupled monomeric actin to divinylsulphone-activated agarose (Mini-Leak) to isolate G-ABPs in human skeletal muscle. Eluted proteins were analyzed by two‐dimensional gel electrophoresis (2-DE), which shows that some proteins are selectively retained. Deoxyribonuclease I (DNase I) is known to bind residues at the "pointed end" of actin (subdomains 2 and 4) with a high affinity. When DNase I is bound to the actin Mini‐Leak before applying the skeletal muscle extract, the 2-DE gels of the eluted proteins reveals differences when compared to gels of proteins eluted from actin-Mini-Leak and DNase I-Mini‐Leak affinity columns. This strategy should detect ABPs which bind to sites other than the DNase I-binding site and some may prove to be novel.
Publication Type: Journal Article
Source of Publication: Electrophoresis, 18(7), p. 1079-1085
Publisher: Wiley-VCH Verlag GmbH & Co KGaA
Place of Publication: Germany
ISSN: 1522-2683
Field of Research (FOR): 060101 Analytical Biochemistry
060109 Proteomics and Intermolecular Interactions (excl. Medical Proteomics)
060199 Biochemistry and Cell Biology not elsewhere classified
Peer Reviewed: Yes
HERDC Category Description: C1 Refereed Article in a Scholarly Journal
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School of Science and Technology

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