Preliminary crystallographic analysis of the complex of the human GTPase RhoA with the DH/PH tandem of PDZ-RhoGEF.
PDZ-containing RhoGEF (PDZ-RhoGEF) is a multidomain protein composed of 1522 amino acids that belongs to the guanine nucleotide exchange factors family (GEF) active on Rho GTPases. It is highly specific for RhoA and is thought to transduce signals from Galpha(12/13)-coupled receptors to the RhoA-dependent regulatory cascades. The protein shows high sequence homology to LARG, p115-RhoGEF and Drosophila DRhoGEF2. The exchange reaction is catalyzed by a DH domain, which is directly downstream of a PH domain in all known Rho-specific GEFs. The DH/PH tandem of PDZ-RhoGEF and C-terminally truncated RhoA were overexpressed in Escherichia coli as TEV protease-cleavable fusion proteins containing GST and a hexahistidine tag at the N-termini, respectively. The nucleotide-free DH/PH-RhoA complex was purified by gel filtration and crystallized. The crystals belong to space group P2(1), with unit-cell parameters a = 88.6, b = 119.0, c = 91.5 A, beta = 114.7 degrees.
Oleksy
Arkadiusz
A
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, VA 22908-0736, USA.
Barton
Holly
H
Devedjiev
Yancho
Y
Purdy
Michael
M
Derewenda
Urszula
U
Otlewski
Jacek
J
Derewenda
Zygmunt S
ZS
D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D020662:Guanine Nucleotide Exchange Factors; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D046911:Macromolecular Substances; D011485:Protein Binding; D017470:Receptors, Glutamate / Q000737:chemistry* / Q000378:metabolism; D064067:Rho Guanine Nucleotide Exchange Factors; D062367:ortho-Aminobenzoates / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000737:chemistry* / Q000378:metabolism
C577234:ARHGEF11 protein, human; D020662:Guanine Nucleotide Exchange Factors; D046911:Macromolecular Substances; D017470:Receptors, Glutamate; D064067:Rho Guanine Nucleotide Exchange Factors; D062367:ortho-Aminobenzoates; C066737:RHOA protein, human; C063981:2'(3')-O-(N-methyl)anthraniloylguanosine 5'-triphosphate; D006160:Guanosine Triphosphate; D020742:rhoA GTP-Binding Protein