Mapping the cytochrome c(553) interacting site using H-1 and N-15 NMR

Cytochrome c(553) is the electron transfer partner of formate dehydrogenase and of [Fe]-hydrogenase, two metalloenzymes essential in the metabolism of sulfate reducing bacteria. These two enzymes contain a 'ferredoxin-like' d omain,which presents 30% identity with Desulfovibrio desulfuricans Norway f erredoxin I. This,vas chosen as a model for the 'ferredoxin-like' domain in volved in the electron transfer reaction with cytochrome c(553), 1D NMR tit ration of complex formation gave us the stoichiometry (1:1) and the dissoci ation constant of the complex (K-d similar to 3 x 10(-6) M). 2D heteronucle ar NMR experiments were performed to analyze the H-1 and N-15 chemical shif t variations that are induced hy the protein-protein recognition. This is t he first mapping of the interaction site on a c-type cytochrome, using hete ronuclear NMR. (C) 1999 Federation of European Biochemical Societies.