YEAST RAS2 MUTATIONS MODULATING THE RAS-GUANINE EXCHANGE FACTOR INTERACTION

We have used a two-hybrid approach to test various forms of Saccharomy ces cerevisiae Ras2p for their ability to interact with the human guan ine nucleotide exchange factor HGRF55. We have previously shown that a strong two-hybrid interaction is found between the HGRF55p and the do minant negative Ras2p(G22A) form of ras [Camus et al, (1995) Oncogene 11, 951-959], We show here that the substitution N123I which weakens t he guanine nucleotide binding also promotes ras-GEF interaction, We de monstrate that the R80D substitution alone completely abolishes the in teraction of Ras2p(G22A) with GEF, whereas substitutions at positions 81, 82 and 73 have only small effects, Since residue 73 is involved in the response of ras to GEF, we propose that it plays a role in the co nformational change induced by the GEF rather than in its binding, Tho se results emphasize the role of the alpha 2 helix of the switch II re gion in the recognition of the GEF family. (C) 1997 Federation of Euro pean Biochemical Societies.