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J. Biol. Chem. 285 (27): 21070-21081

© 2010 by The American Society for Biochemistry and Molecular Biology, Inc.

Activated RhoA Binds to the Pleckstrin Homology (PH) Domain of PDZ-RhoGEF, a Potential Site for Autoregulation*Formula {diamondsuit}

Zhe Chen{ddagger}12, Frank Medina{ddagger}1, Mu-ya Liu{ddagger}, Celestine Thomas§, Stephen R. Sprang§, , and Paul C. Sternweis{ddagger}3

From the {ddagger}Department of Pharmacology, The University of Texas Southwestern Medical Center, Dallas, Texas 75390 and
the §Center for Biomolecular Structure and Dynamics, Division of Biological Sciences, University of Montana, Missoula, Montana 59812

ABSTRACT Back to Top

Abstract: Guanine nucleotide exchange factors (GEFs) catalyze exchange of GDP for GTP by stabilizing the nucleotide-free state of the small GTPases through their Dbl homology/pleckstrin homology (DH·PH) domains. Unconventionally, PDZ-RhoGEF (PRG), a member of the RGS-RhoGEFs, binds tightly to both nucleotide-free and activated RhoA (RhoA·GTP). We have characterized the interaction between PRG and activated RhoA and determined the structure of the PRG-DH·PH-RhoA·GTP{gamma}S (guanosine 5'-O-[{gamma}-thio]triphosphate) complex. The interface bears striking similarity to a GTPase-effector interface and involves the switch regions in RhoA and a hydrophobic patch in PRG-PH that is conserved among all Lbc RhoGEFs. The two surfaces that bind activated and nucleotide-free RhoA on PRG-DH·PH do not overlap, and a ternary complex of PRG-DH·PH bound to both forms of RhoA can be isolated by size-exclusion chromatography. This novel interaction between activated RhoA and PH could play a key role in regulation of RhoGEF activity in vivo.

Key Words: G Proteins • Protein Structure • Protein-Protein Interactions • Signal Transduction • X-ray Crystallography

Received for publication March 11, 2010. Revision received April 15, 2010.

FOOTNOTES Back to Top

1 Both authors contributed equally to this work.

2 To whom correspondence may be addressed. E-mail: Zhe.Chen{at}utsouthwestern.edu.

3 To whom correspondence may be addressed: Dept. of Pharmacology, The University of Texas Southwestern Medical Center, 6001 Forest Park Rd., Dallas, TX 75390. Fax: 214-645-6151; E-mail: Paul.Sternweis{at}utsouthwestern.edu.

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