Identification of amino acid residues required for Ras p21 target activation.
The Krev-1 gene has been shown to suppress ras-mediated transformation in vitro. Both ras and Krev-1 proteins have identical effector domains (ras residues 32 to 40), which are required for biological activity and for the interaction of Ras p21 with Ras GTPase-activating protein (GAP). In this study, five amino acid residues flanking the ras effector domain, which are not conserved with the Krev-1 protein, were shown to be required for normal protein-protein interactions and biological activity. The substitution of Krev-1 p21 residues 26, 27, 30, 31, and 45 with the corresponding amino acid residues from Ras p21 resulted in a Krev-1 protein which had ras function in both mammalian and yeast biological assays. Replacement of these residues in Ras p21 with the corresponding Krev-1 p21 amino acids resulted in ras proteins which were impaired biologically or reduced in their affinity for in vitro GAP binding. Evaluation of these mutant ras proteins have implications for Ras p21-GAP interactions in vivo.
Marshall
M S
MS
Department of Cancer Research, Merck Sharp and Dohme Research Laboratories, West Point, Pennsylvania 19486.
Davis
L J
LJ
Keys
R D
RD
Mosser
S D
SD
Hill
W S
WS
Scolnick
E M
EM
Gibbs
J B
JB
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D004261:DNA Replication; D004274:DNA, Recombinant / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics*; D011905:Genes, ras; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015689:Oncogene Protein p21(ras) / Q000235:genetics*; D011518:Proto-Oncogene Proteins / Q000235:genetics; D011519:Proto-Oncogenes; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D013489:Suppression, Genetic; D013936:Thymidine / Q000378:metabolism; D014162:Transfection; D020668:rap GTP-Binding Proteins
D004274:DNA, Recombinant; D011518:Proto-Oncogene Proteins; D019204:GTP-Binding Proteins; D015689:Oncogene Protein p21(ras); D020668:rap GTP-Binding Proteins; D013936:Thymidine