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Identifying competitive protein antagonists for F-actin with reverse-phase high-performance liquid chromatography.

ABSTRACT: F-actin binding constants are traditionally determined by centrifugal cosedimentation with actin microfilaments, where bound protein is separated from actin with SDS-PAGE and quantitated using densitometry. Here, we demonstrate that UV quantitation of reverse-phase HPLC-separated proteins provides increased accuracy and sensitivity, can be fully automated, and allows one to perform F-actin competition assays on similar sized proteins.

SUBMITTER: Brown JW

PROVIDER: S-EPMC2812587 | biostudies-literature | 2010 Mar

REPOSITORIES: biostudies-literature

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Identifying competitive protein antagonists for F-actin with reverse-phase high-performance liquid chromatography.

Brown Jeffrey W JW McKnight C James CJ

Analytical biochemistry 20091120 1

F-actin binding constants are traditionally determined by centrifugal cosedimentation with actin microfilaments, where bound protein is separated from actin with SDS-PAGE and quantitated using densitometry. Here, we demonstrate that UV quantitation of reverse-phase HPLC-separated proteins provides increased accuracy and sensitivity, can be fully automated, and allows one to perform F-actin competition assays on similar sized proteins. ...[more]

PMID: 19932072

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