Electric-field-induced redox potential shifts of tetraheme cytochromes c3 immobilized on self-assembled monolayers: Surface-enhanced resonance raman spectroscopy and simulation studies

Abstract

The tetraheme protein cytochrome c3 (Cyt-c3) from Desulfovibrio gigas, immobilized on a self-assembled monolayer (SAM) of 11-mercaptoundecanoic acid, is studied by theoretical and spectroscopic methods. Molecular dynamics simulations indicate that the protein docks to the negatively charged SAM via its lysine-rich domain around the exposed heme IV. Complex formation is associated with only little protein structural perturbations. This finding is in line with the resonance Raman and surface-enhanced resonance Raman (SERR) spectroscopic results that indicate essentially the same heme pocket structures for the protein in solution and adsorbed on SAM-coated Ag electrodes. Electron- and proton-binding equilibrium calculations reveal substantial negative shifts of the redox potentials compared to the protein in solution. The magnitude of these shifts decreases in the order heme IV (-161 mV) > heme III (-73 mV) > heme II (-57 mV) > heme I (-26 mV), resulting in a change of the order of reduction. These shifts originate from the distance-dependent electrostatic interactions between the SAM headgroups and the individual hemes, leading to a stabilization of the oxidized forms. The results of the potential-dependent SERR spectroscopic analyses are consistent with the theoretical predictions and afford redox potential shifts of -160 mV (heme IV), -90 mV (heme III), -70 mV (heme II), and +20 mV (heme I) relative to the experimental redox potentials for Cyt-c3 in solution. SERR spectroscopic experiments reveal electric-field-induced changes of the redox potentials also for the structurally very similar Cyt-c3 from Desulfovibrio vulgaris, although the shifts are somewhat smaller compared to Cyt-c3 from D. gigas. This study suggests that electric-field-induced redox potential shifts may also occur upon binding to biomembranes or partner proteins and thus may affect biological electron transfer processes. © 2005 by the Biophysical Society.