Role of phenylalanine-82 in yeast iso-1-cytochrome c and remote conformational changes induced by a serine residue at this position

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Abstract
A three-dimensional structural analysis of the reduced form of the Ser-82 mutant protein of yeast iso-1 -cytochrome c has been completed to 2.8-.ANG. resolution. Replacement of Phe-82 with a serine residue results in conformational changes both near and remote from the mutation site. Those groups undergoing positional shifts near Ser-82 include Arg-13, Gly-83 and -84, and the CBB methyl of the heme group. Remote shifts are centered about the propionate of pyrrole ring A and principally involve Asn-52, Trp-59, and an internally buried water molecule, WAT-166. Placement of a serine side chain at position 82 also leads to the formation of a large solvent channel which substantially increases the solvent accessibility of the heme group. This would appear to account for the much lower reduction potential observed for this protein. The detrimental effect of Ser-82 on both the steady-state activity and the rate of electron transfer in complexation with cytochrome c peroxidase can also be interpreted in terms of the modified character of the region about the mutation site. The remote conformational changes observed appear to represent the equivalent of the initial conformational changes occurring as yeast iso-1 -cytochrome c is converted to the fully oxidized state during an electron-transfer event. These results agree well with the proposal [Moore, G. R. (1983) FEBS Lett. 161, 171-175] that the trigger for conformational changes between oxidation states resides in the nature of the interactions between the heme iron atom and the pyrrole ring A propionate group. Overall, our results support suggestions that Phe-82 in the wild-type protein has at least three roles. These include the following: limiting solvent accessibility to the heme, thereby regulating the reduction potential of this protein dielectrically; facilitating the rate of electron transfer by providing the optimal medium along the transfer route; forming contact face interactions with redox partners to assist in th formation of the productive electron-transfer complex.