Identification of the guanine nucleotide dissociation stimulator for Ral as a putative effector molecule of R-ras, H-ras, K-ras, and Rap.
To identify proteins that bind to the Ras-related protein R-ras we performed a yeast two-hybrid cDNA library screen. Several clones were obtained encoding the C-terminal region of the guanine nucleotide dissociation stimulator for Ral (RalGDS). The R-ras-binding domain of RalGDS (RalGDS-RBD) is distinct from the conserved catalytic exchange factor regions. Using the two-hybrid system, we show that RalGDS-RBD interacts with H-ras, K-ras, and Rap, and with active but not with inactive point mutants of these Ras-like GTPases. Moreover, using purified proteins, we demonstrate the direct GTP-dependent interaction of the Ras-like GTPases with RalGDS-RBD and full-length RalGDS in vitro. Furthermore, we show that RalGDS-RBD and the Ras-binding domain of Raf-1 compete for binding to the Ras-like GTPases. These data indicate that RalGDS is a putative effector molecule for R-ras, H-ras, K-ras, and Rap.
Spaargaren
M
M
ONYX Pharmaceuticals, Richmond, CA 94806.
Bischoff
J R
JR
D000595:Amino Acid Sequence; D000818:Animals; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D020725:ral GTP-Binding Proteins; D020726:ral Guanine Nucleotide Exchange Factor; D020668:rap GTP-Binding Proteins; D018631:ras Proteins / Q000378:metabolism*
D011518:Proto-Oncogene Proteins; D020726:ral Guanine Nucleotide Exchange Factor; D006160:Guanosine Triphosphate; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; C085929:RRAS protein, human; C513254:Rras protein, mouse; D020725:ral GTP-Binding Proteins; D020668:rap GTP-Binding Proteins; D018631:ras Proteins