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Determining Rho GTPase activity by an affinity-precipitation assay

Research output: Chapter in Book/Report/Conference proceedingChapter

Original languageEnglish
Title of host publicationAdhesion Protein Protocols
EditorsAmanda S. Coutts
Place of PublicationNew York
PublisherHumana Press
Pages191-202
Number of pages12
VolumeN/A
Edition3
ISBN (Print)9781627035378
DOIs
Published2013

Publication series

NameMethods in Molecular Biology
PublisherHumana Press
Volume1046
ISSN (Print)1064-3745

King's Authors

Abstract

The Rho GTPases are members of the Ras superfamily of GTPases that are pivotal regulators of the actin cytoskeleton. They also contribute to other cellular processes such as gene transcription, cell polarity, microtubule dynamics, cell cycle progression and vesicle trafficking. Most Rho GTPases act as molecular switches cycling between an "active" GTP-bound form and an "inactive" GDP-bound form. Hence, to elucidate the mechanisms by which Rho GTPases regulate cellular responses, an important parameter to determine is the GTP-loading of each Rho family member in cells under different conditions. Here we describe a biochemical technique to assess this based on affinity-precipitation of the GTP-bound form from whole cell lysates.

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