Getting to heme. A system for the incorporation of unnatural amino acid into cytochrome c3 by site‐specific viologen modification has been established. A viologen was linked to the side chain of the unnatural amino acid, and cytochrome c3 accepted an electron through the linked viologen.

Cytochrome c3 from Desulfovibrio vulgaris (Miyazaki) F is an electron-transfer protein between hydrogenase and ferredoxin and other types of cytochromes. In this study, an N-terminus of the cytochrome c3 has been identified as an N-methylated alanine residue by cleavage with peptidase, liquid chromatography/mass spectrometry (LC/MS) and nuclear magnetic resonance (NMR) analysis. Heterologous expression of the cytochrome c3 with Shewanella oneidensis TSP-C is not methylated, this indicate that the N-methylation occur in vivo in D. vulgaris.

No abstract

Cytochrome c 3 from Desulfovibrio vulgaris (Miyazaki) is an electron transfer protein containing four hemes per molecule. Its physiological electron transfer partner is the hydrogenase which catalyzes reversible oxidation of hydrogen. The complex formation between cytochrome c 3 and hydrogenase is caused by electrostatic interaction, because cytochrome c 3 is a basic protein and hydrogenase is an acidic protein. As cytochrome c 3 has 20 lysine residues among 108 amino acids, the positive charges of some lysine residues may play an important role in the interaction with hydrogenase. To clarify the role of positive charge of lysine residue, the positive charge was changed to neutral or negative charge using chemical modification and site-directed mutagenesis. When the positive charges around heme IV were changed, the hydrogen evolution rate with hydrogenase decreased. The affinity between hydrogenase and mutated cytochrome c 3 (K57Q, K57E, K72Q, K94Q, K94E) were not affected. On the other hand, the affinity of K72E cytochrome c 3 for hydrogenase was very low. These results suggest that the positive charge around heme IV plays an important role in the electrostatic interaction with hydrogenase in hydrogen evolution.