Discriminatory residues in Ras and Rap for guanine nucleotide exchange factor recognition

The inability of the S17N mutant of Rap1A to sequester the catalytic domain of the Rap guanine nucleotide exchange factor C3G (van den Berghe, N., Coo l, R, H., Horn, G., and Wittinghofer, A. (1997) Oncogene 15, 845-850) promp ted us to study possible fundamental differences in the way Rap1 interacts with C3G compared with the interaction of Ras with the catalytic domain of the mouse Ras guanine nucleotide exchange factor Cdc25(Mm), A variety of mu tants in both Ras and Rap1A were designed, and both the C3G and Cdc25(Mm) c atalyzed release of guanine nucleotide from these mutants was studied. In a ddition, we could identify regions in Rap2A that are responsible for the la ck of recognition by C3G and induce high C3G activity by replacement of the se residues with the corresponding Rap1A residues. The different Ras and Ra p mutants showed that many residues were equally important for both C3G and Cdc25(Mm), suggesting that they interact similarly with their substrates, However, several residues were also identified to be important for the exch ange reaction with only C3G (Leu(70)) or only Cdc25(Mm) (Gln(61) and Tyr(40 )). These results are discussed in the light of the structure of the Ras-So s complex and suggest that some important differences in the interaction of Rap1 with C3G and Ras with Cdc25(Mm) indeed exist and that marker residues have been identified for the different structural requirements.