Influence of guanine nucleotides on complex formation between Ras and CDC25 proteins.

摘要: The Saccharomyces cerevisiae CDC25 gene and closely homologous genes in other eukaryotes encode guanine nucleotide exchange factors for Ras proteins. We have determined the minimal region of budding yeast capable activity vivo. required full biological is approximately 450 residues contains two segments to proteins: one found both Ras-specific more distant Bud5 Lte1 proteins, a smaller segment 48 amino acids only factors. When expressed Escherichia coli as fusion protein, this was be potent catalyst GDP-GTP on Ras2 well human p21H-ras but inactive promoting Ras-related proteins Ypt1 Rsr1. protein catalyzed replacement GDP-bound with GTP (activation) efficiently than that reverse reaction GDP (deactivation), consistent prior genetic analysis which indicated positive role activation Ras. To directly study physical interaction we developed protein-protein binding assay. binds tightly absence nucleotides. This higher affinity nucleotide-free form either GDP- or GTP-bound suggests catalyzes nucleotides bound by stabilization transitory state.