Mutations at position 1122 in the catalytic domain of the mouse ras-specific guanine nucleotide exchange factor CDC25(Mm) originate both loss-of-function and gain-of-function proteins

role of two residues within the catalytic domain of CDC25(Mm), a mouse ras- specific guanine nucleotide exchange factor (GEF), was investigated by site -directed mutagenesis, The function of the mutant proteins was tested in vi vo in both a Saccharomyces cerevisiae cdc25 complementation assay and in a mammalian fos-luciferase assay, and in in vitro assays on human and yeast R as proteins. Mutants CDC25(MmE1048K) and CDC25(MmS1122V) were shown to be ( partly) inactive proteins, similar to their yeast homologs, Mutant CDC25(Mm S1122A) showed higher nucleotide exchange activity than the wild type prote in on the basis of both in vitro and in vivo assays. Thus, alanine and vali ne substitutions at position 1122 within the GEF catalytic domain originate mutations with opposite biological properties, indicating an important rol e for position 1122 in GEF function. (C) 1998 Federation of European Bioche mical Societies.