c-Raf-1 RBD associates with a subset of active v-H-ras

M. Fridman, F. Walker, B. Catimel, T. Domagala, E. Nice, A. Burgess

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4 Citations (Scopus)

Abstract

Mutational analysis of the cRaf-1 Ras binding domain (RBD) identified several point mutants with elevated Ras binding. Detailed examination of the binding kinetics of one mutant (A85K) suggests that it associates with a greater range of isomeric conformers of v-H-Ras than wt-RBD. At limiting v-H-Ras concentrations, saturation binding to A85K-RBD is higher than to wt-RBD. Notably, in assay systems where the RBD concentration is limiting, no difference exists between wt-RBD and A85K-RBD saturation levels in the presence of a sufficiently large molar excess of Ras. The inability of wt-RBD to saturate all bindable Ras/GTP (defined by its binding to A85K-RBD) suggests that Ras/GTP exists as several isoforms and that only a minority of these isoforms are capable of associating with wt-RBD. These findings provide the first experimental evidence in support of functionally distinct Ras/GTP isoforms. We also describe a novel analysis of such isoforms.

Original languageEnglish
Pages (from-to)15603-15611
Number of pages9
JournalBiochemistry
Volume39
Issue number50
DOIs
Publication statusPublished - 19 Dec 2000
Externally publishedYes

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