Resonance Raman fingerprinting of multiheme cytochromes from the cytochrome c₃ family

Resonance Raman (RR) spectroscopy was used to investigate conformational characteristics of the hemes of several ferricytochromes of the cytochrome c₃ family, electron transfer proteins isolated from the periplasm and membranes of sulfate-reducing bacteria. Our analysis concentrated on the low-frequency region of the RR spectra, a fingerprint region that includes vibrations for heme-protein C-S bonds [ν(C_aS)]. It has been proposed that these bonds are directly involved in the electron transfer process. The three groups of tetraheme cytochrome c₃, analyzed, namely Type I cytochrome c₃ (TpIc₃s), Type II cytochrome c₃ (TpIIc₃s) and Desulfomicrobium cytochromes c ₃, display different frequency separations for the two ν(C _aS) lines that are similar among members of each group. These spectral differences correlate with differences in protein structure observed among the three groups of cytochromes c₃. Two larger cytochromes of the cytochrome c₃ family display RR spectral characteristics for the ν(C_aS) lines that are closer to TpIIc₃ than to TpIc₃. Two other multiheme cytochromes from Desulfovibrio that do not belong to the cytochrome c₃ family display ν(C_aS) lines with reverse relative areas in comparison with the latter family. This RR study shows that the small differences in protein structure observed among these cytochrome c₃ correlate to differences on the heme-protein bonds, which are likely to have an impact upon the protein function, making RR spectroscopy a sensitive and useful tool for characterizing these cytochromes.