Influence of the Interdomain Interface on Structural and Redox Properties of Multiheme Proteins.

Multiheme proteins are important in energy conversion and biogeochemical cycles of nitrogen and sulfur. A diheme cytochrome () was used as a model to elucidate roles of the interdomain interface on properties of iron centers in its hemes A and B. Isolated monoheme domains -A and -B, together with the full-length diheme and its Met-to-His ligand variants, were characterized by a variety of spectroscopic and stability measurements. In both isolated domains, the heme iron is Met/His-ligated at pH 5.0, as in the full-length , but becomes His/His-ligated in -B at higher pH. Intradomain contacts in -A are minimally affected by the separation of -A and -B domains, and isolated -A is folded. In contrast, the isolated -B is partially unfolded, and the interface with -A guides folding of this domain. The -A and -B domains have the propensity to interact even without the polypeptide linker. Thermodynamic cycles have revealed properties of monomeric folded isolated domains, suggesting that ferrous (Fe), but not ferric (Fe) -A and -B, is stabilized by the interface. This study illustrates the effects of the interface on tuning structural and redox properties of multiheme proteins and enriches our understanding of redox-dependent complexation.