Kazanskaya I, Lexa D, Bruschi M, Chottard G
Chimie des Métaux de Transition, Université Pierre et Marie Curie, Paris, France.
Biochemistry. 1996 Oct 15;35(41):13411-8. doi: 10.1021/bi9608424.
Electron transfer in tetrahemic cytochromes c3 from Desulfovibrio vulgaris Hildenborough (D.v.H.) and Desulfovibrio desulfuricans Norway (D.d.N.) strains has been investigated by thin layer spectroelectrochemistry with visible absorption, CD, and resonance Raman (RR) monitoring. The observed splitting of the isosbestic point in the Soret absorption band indicates that the electron transfer from the (FeIII)4 state to the (FeII)4 state proceeds via an intermediate species, which corresponds to 25 and 50% reduction for the D.v.H. cyt.c3 and the D.d.N. cyt.c3, respectively. For the latter, a specific CD signal is observed at half-reduction. RR monitoring of the redox process does not reveal multiple splitting of the high-frequency RR bands, at variance with previously published results on the enzymatic reduction of cyt.c3 from Desulfovibrio vulgaris Miyazaki, a cytochrome highly homologous to D.v.H. cyt.c3 [Verma, A.L., Kimura, A., Nakamura, A., Yagi, T., Inoguchi, H., & Kitagawa, T. (1988) J. Am. Chem. Soc. 110, 6617-6623]. The low-frequency RR spectra of the intermediate species differ significantly from the ones calculated from a linear combination of the all-ferric and all-ferrous states, for the same reduction ratio. Frequency shifts of the bending modes of the cysteine and propionate heme substituents are observed, as well as changes specific to each cytochrome; most notable is the activation of two torsional modes in the case of D.d.N. cyt.c3. Comparison of the results obtained for the two cytochromes leads to the conclusion that reduction of heme IV triggers the observed conformational change. This conclusion is supported by the spectroelectrochemical investigation of the mutant D.v.H. cyt.c3 H25M, in which the sixth ligand of heme III, histidine, is replaced by a methionine.
利用薄层光谱电化学技术，通过可见吸收光谱、圆二色光谱（CD）和共振拉曼光谱（RR）监测，对来自希登伯勒脱硫弧菌（D.v.H.）和挪威脱硫脱硫弧菌（D.d.N.）菌株的四血红素细胞色素c3中的电子转移进行了研究。在Soret吸收带中观察到的等吸收点分裂表明，从（FeIII）4态到（FeII）4态的电子转移是通过一种中间物种进行的，对于D.v.H.细胞色素c3和D.d.N.细胞色素c3，该中间物种分别对应25%和50%的还原率。对于后者，在半还原时观察到一个特定的CD信号。氧化还原过程的RR监测未发现高频RR带的多重分裂，这与先前发表的关于宫崎脱硫弧菌细胞色素c3酶促还原的结果不同，宫崎脱硫弧菌细胞色素c3是一种与D.v.H.细胞色素c3高度同源的细胞色素[Verma, A.L., Kimura, A., Nakamura, A., Yagi, T., Inoguchi, H., & Kitagawa, T. (1988) J. Am. Chem. Soc. 110, 6617 - 6623]。对于相同的还原率，中间物种的低频RR光谱与由全铁态和全亚铁态的线性组合计算得到的光谱有显著差异。观察到半胱氨酸和丙酸血红素取代基弯曲模式的频率位移，以及每种细胞色素特有的变化；最值得注意的是，在D.d.N.细胞色素c3的情况下，两种扭转模式被激活。对这两种细胞色素所得结果的比较得出结论，血红素IV的还原引发了观察到的构象变化。这一结论得到了突变体D.v.H.细胞色素c3 H25M的光谱电化学研究的支持，在该突变体中，血红素III的第六个配体组氨酸被甲硫氨酸取代。
