Expression, Purification, and Characterization of an Enzymatically Active Truncated Human Rho-Kinase I (ROCK I) Domain Expressed in Sf-9 Insect Cells

Author(s): Sanjay S. Khandekar, Tracey Yi, Ed Dul, Lois L. Wrigh, Susan Chen, Gilbert F. Scott, Gary K. Smith, Dennis Lee, Erding Hu, Robert B. Kirkpatrick

Journal Name: Protein & Peptide Letters

Volume 13 , Issue 4 , 2006

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Abstract:

Rho Kinase I (ROCK I) is a serine/threonine kinase that is involved in diverse cellular signaling. To further understand the physiological role of ROCK I and to identify and develop potent and selective inhibitors of ROCK I, we have overexpressed and purified a constitutively active dimeric human ROCK I (3-543) kinase domain using the Sf9- baculovirus expression system. In addition, using a limited proteolysis technique, we have identified a minimal functional subdomain of ROCK I that can be used in crystallization studies. The availability of multimilligram amounts of purified and well characterized functional human ROCK I kinase domains will be useful in screening and structural studies.

Keywords: ROCK I, expression, kinetics, inhibition, kinase domain

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Article Details

VOLUME: 13
ISSUE: 4
Year: 2006
Page: [369 - 376]
Pages: 8
DOI: 10.2174/092986606775974357
Price: $65

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