Resonance Raman investigation of dithionite-reduced cobalamin

Absorption (Abs) and resonance Raman (rR) spectroscopy along with density functional theory (DFT) have been applied to investigate the derivatives of vitamin B-12 (or cobalamins = Cbls). Cbls are cobalt-containing corrin complexes, vibrations of which can be probed using rR spectroscopy. The oxidation state of Co ion in Cbls can range between Co3+, Co2+, and Co1+, and axially, cobalt can be coordinated by variety of ligands. Recent spectroscopic study revealed that Co(2+)Cbl reduced by dithionite (DTH) exhibited a unique property distinct from other Co(2+)Cbls. Accordingly, in this study, rR spectra of Co(2+)Cbls reduced by DTH were compared with these reduced by dithiothreitol (DTT). The DTT-reduced Cbl gave ordinary rR spectrum at pH 3 and 8. On the other hand, the DTH-reduced Cbl gave different rR spectra at pH 3 and 8 and contained a new rR band at 987 cm(-1), which is absent with the DTT-reduced one. The alpha and gamma Abs bands of DTH-reduced Cbls were also shifted to shorter wavelengths and exhibited broadening. To understand these rR spectra, geometry optimization and frequency calculations using DFT have been carried out for different structural models of Cbls and compared with experiment. Based on current spectroscopic studies and computation, we deduced that the DTH-reduced Cbl has SO2-. radical anion at an axial coordination position of Co2+ and that the pi pi* transition of corrin ring is partially mixed with pi bond of SO2-. radical anion, giving rise to the S=O stretching rR band in resonance with the gamma Abs band.