Abstract
The tendency of the periplasmic redox proteins in acidophiles to have more positive redox potentials (E(m)) than their homologous counterparts in neutrophiles suggests an adaptation to acidic pH at protein level, since thermodynamics of electron transfer processes are also affected by acidic pH. Since this conclusion is mainly based on the electrochemical characterization of redox proteins from extreme acidophiles of the genus Acidithiobacillus, we aimed to characterize three recombinant redox proteins of the more moderate acidophile Ferrovum sp. PN-J47-F6. We applied protein film voltammetry and linear sweep voltammetry coupled to UV/Vis spectroscopy to characterize the redox behavior of HiPIP-41, CytC-18, and CytC-78, respectively. The E(m)-values of HiPIP-41 (571 ± 16 mV), CytC-18 (276 ± 8 mV, 416 ± 2 mV), and CytC-78 (308 ± 7 mV, 399 ± 7 mV) were indeed more positive than those of homologous redox proteins in neutrophiles. Moreover, our findings suggest that the adaptation of redox proteins with respect to their E(m) occurs more gradually in response to the pH, since there are also differences between moderate and more extreme acidophiles. In order to address structure function correlations in these redox proteins with respect to structural features affecting the E(m), we conducted a comparative structural analysis of the Ferrovum-derived redox proteins and homologs of Acidithiobacillus spp. and neutrophilic proteobacteria. Hydrophobic contacts in the redox cofactor binding pockets resulting in a low solvent accessibility appear to be the major factor contributing to the more positive E(m)-values in acidophile-derived redox proteins. While additional cysteines in HiPIPs of acidophiles might increase the effective shielding of the [4Fe-4S]-cofactor, the tight shielding of the heme centers in acidophile-derived cytochromes is achieved by a drastic increase in hydrophobic contacts (A.f. Cyc(41)), and by a larger fraction of aromatic residues in the binding pockets (CytC-18, CytC-78).