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Quantitative structure-activity analysis correlating Ras/Raf interaction in vitro to Raf activation in vivo.

C Block·R Janknecht·C Herrmann·N Nassar·A Wittinghofer
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Abstract

Binding of Ras to c-Raf-1 is a pivotal step of many mitogenic signalling pathways. Based on the recent crystal structure of the complex of Rap1A with the Ras-binding domain of Raf, mutations were introduced in c-Raf-1 and their effects on Ras/Raf binding affinity in vitro and Ras/Raf regulated gene expression in vivo were analysed. Our data reveal an empirical semilogarithmic correlation between dissociation constants and Raf-induced gene activity. The functional epitope that primarily determines binding affinity consists of residues Gln 66, Lys 84 and Arg 89 in Raf. This quantitative structure-activity investigation may provide a general approach to correlate structure-guided biochemical analysis with biological function of protein-protein interactions.

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Amino Acid SequenceAnimalsBinding SitesCircular DichroismCrystallography, X-RayGTP-Binding ProteinsGuanylyl ImidodiphosphateHumansKineticsModels, MolecularMolecular Sequence DataMutagenesis, Site-DirectedPoint MutationProtein ConformationProtein Serine-Threonine KinasesProto-Oncogene ProteinsProto-Oncogene Proteins c-rafSequence Homology, Amino AcidSignal Transductionrap GTP-Binding ProteinsComparative StudyJournal ArticleResearch Support, Non-U.S. Gov't

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C Block, R Janknecht, C Herrmann, N Nassar, A Wittinghofer. 1996. Quantitative structure-activity analysis correlating Ras/Raf interaction in vitro to Raf activation in vivo.. https://doi.org/10.1038/nsb0396-244

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