Electron transfer properties of the soluble Cyt-c domain from Rhodothermus marinus oxygen reductase

MOLINAS, M.F.; CAPDEVILLA, D.; DE CANDIA, A.G.; RODRÍGUEZ, J.B.; TODOROVIC, S.; MARTI, M. A. AND MURGIDA, D. H

Salta

Encuentro; 3rd Latin American Protein Society Meeting; 2010

Latinamerican Protein Society

Heme/copper oxygen reductases (ORs) constitute a superfamily of membrane-bound enzymes that catalyse the reduction of dioxygen to water as a terminal step in the respiratory chains of aerobic organisms. We studied the electron transfer (ET) properties of Rhodothermus marinus caa (OR) or cytochrome-c domain (Rm-ccd) and its putative partner, a high potential iron-sulfur protein (HiPIP). We found that, according to circular dichroism and UV-vis titrations Rm-ccd exhibits a thermal, pH and GdnHCl stability superior to other cytochromes. Using SERR, TR-SERR, cyclic voltammetry and molecular dynamics (MD) simulation we determined that Rm-ccd can be immobilized on neutral and charged biomimetic films, retaining its native structure. By analyzing relative SERR intensities of modes of different symmetry it is inferred that the adsorbed protein is well oriented and it also shows the same redox potential as in solution. Measured ET rates are unusually fast compared to soluble cytochromes, and MD simulations shows that the protein is adsorbed trough an specific patch that leaves the heme edge in very close proximity to the surface, thus providing very high electronic coupling. HiPIP was immobilized in films of the same composition and has a very similar redox potential, indicating that the reaction has a low ΔG value. In summary, the results suggest that Rm-ccd has been evolutionary optimized for fast ET to a greater extent than usual both in terms of low reorganization energy and high electronic coupling.