A single mutation in the heme 4 environment of Desulfovibrio desulfuricans Norway cytochrome c(3) (M-r 26,000) greatly affects the molecule reactivity

The gene encoding Desulfovibrio desulfuricans Norway cytochrome c(3) (M-r 26,000), a dimeric octaheme cytochrome belonging to the polyheme cytochrome c(3) superfamily, has been cloned and successfully expressed in another sulfate reducing bacteria, D. desulfuricans G201. The gene, named cycD, is monocistronic and encodes a cytochrome precursor of 135 amino acids with an extension at the NH2 terminus of 24 amino acids. This extension acts as a signal sequence which allows export across the cytoplasmic membrane into the periplasmic space, Tyrosine 73, which is in a close contact with the histidine sixth axial ligand to the heme 4 iron atom, has been replaced by a glutamate residue using site directed mutagenesis, The cytochrome mutant when expressed in D. desulfuricans G201, is correctly folded and matured, A global increase of the oxidoreduction potentials of about 50 mV is measured for the Y73E cytochrome. The mutation also has a strong influence on the interaction of the cytochrome with its redox partner, the hydrogenase. This suggests, like the tetraheme cytochrome c(3) (M-r 13,000), heme 4 is the interactive heme in the cytochrome-hydrogenase complex and that alteration of the heme 4 environment can greatly affect the electron transfer reaction with its redox partner.