Research On Oxidation-reduction Of Cytochrome C Irradiated By The UV-Vis Light

As a kind of hemoproteins, Cyt c can be irradiated by the UV-Vis light of a specific wavelength to express unfamiliar functions of hemeprotein. UV-Vis absorption spectra, synchronous fluorescence specta and CD spectra were used to investigated the mechanism and effects of different external factors on the photoreduction of Cyt c. Based on the absorbance intensity increase of δ band(317nm) and Q band(520 nm and 549 nm), Cyt c in phosphate-buffered saline under N2 atmosphere was photoreduced to Cyt c-Fe Ⅱ. Considered the temperature, fixed wavelength and p H values, the optimum conditions of photoreduction were irradiated by 410 nm light at 24℃ in alkaline solution. The photoreduction process was affected by adding external amino acids. Met facilitated the photoreduction by irradiation with410 nm, while with the existences of Trp, Tyr and Phe, the process were impeded by the light absorbance abilities of aromatic amino acids. The photoreduction mechanism of Cyt c was intramolecular electron transfer including the formation of porphyrin cation radical as an active intermediate excited by 410 nm light. In addition, results of fluorescence and CD spectra indicated that in the photoreduction process the microenvironment polarity of Trp residue varied and the secondary structure of Cyt c changed with lower α-helix/β-sheet ratio.As an important electron mediator, Cyt c can regulate electron transfer process in cell by the variation of the heme iron atom valence between bivalent and trivalent states. In this work, we found free amino acid played an important role in photoreduction of Cyt c. Applied with UV-Vis absorption and CD spectra, we studied the aromatic amino acids(tryptophan, phenylalanine and tyrosine), aspartic acid,arginine, methionine, cysteine and glutathione impacts on the photoreduction of Cyt c in the crowded environment by adding crowded reagents. The UV-Vis absorptionspectrum data revealed that the amino acids could facilitate the photoreduction process of Cyt c with the following order of promotion effects: Cys> GSH> Phe>Tyr> Asp> Arg> Met> Trp. In addition, the photoreduction extent was much larger in Ficoll70 solution than that in Dextran70. The Cyt c photoreduction process may be radical electron transfer process. By clearing radicals, amino acid could facilitate the photoreduction of Cyt c in simulated physiological environment.