Mutational analysis reveals a single binding interface between RhoA and its effector, PRK1.

Abstract:

:Protein kinase C-related kinases (PRKs) are serine/threonine kinases that are members of the protein kinase C superfamily and can be activated by binding to members of the Rho family of small G proteins via a Rho binding motif known as an HR1 domain. The PRKs contain three tandem HR1 domains at their N-termini. The structure of the HR1a domain from PRK1 in complex with RhoA [Maesaki, R., et al. (1999) Mol. Cell 4, 793-803] identified two potential contact interfaces between the G protein and the HR1a domain. In this work, we have used an alanine scanning mutagenesis approach to identify whether both contact sites are used when the two proteins interact in solution and also whether HR1b, the second HR1 domain from PRK1, plays a role in binding to RhoA. The mutagenesis identified just one contact site as being relevant for binding of RhoA and HR1a in solution, and the HR1b domain was found not to contribute to RhoA binding. The folded state and thermal stability of the HR1a and HR1b domains were also investigated. HR1b was found to be more thermally stable than HR1a, and it is hypothesized that the differences in the biophysical properties of these two domains govern their interaction with small G proteins.

journal_name

Biochemistry

journal_title

Biochemistry

authors

Hutchinson CL,Lowe PN,McLaughlin SH,Mott HR,Owen D

doi

10.1021/bi200039u

subject

Has Abstract

pub_date

2011-04-12 00:00:00

pages

2860-9

issue

14

eissn

0006-2960

issn

1520-4995

journal_volume

50

pub_type

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