Akutsu H, Hazzard J H, Bartsch R G, Cusanovich M A
Department of Biochemistry, University of Arizona, Tucson.
Biochim Biophys Acta. 1992 Dec 7;1140(2):144-56. doi: 10.1016/0005-2728(92)90003-k.
The reduction of the tetraheme cytochrome c3 (from Desulfovibrio vulgaris, strains Miyazaki F and Hildenbourough) by flavin semiquinone and reduced methyl viologen follows a monophasic kinetic profile, even though the four hemes do not have equivalent reduction potentials. Rate constants for reduction of the individual hemes are obtained subsequent to incrementally reducing the cytochrome by phototitration. The dependence of each rate constant on the reduction potential difference between the heme and the reductant can be described by outer sphere electron transfer theroy. Thus, the very low reduction potentials of the cytochrome c3 hemes compensate for the very large solvent accessibility of the hemes. The relative rate constants for electron transfer to the four hemes of cytochrome c3 are consistent with the assignments of reduction potential to hemes previously made by Park et al. (Park, J.-S., Kano, K., Niki, S. and Akutsu, H. (1991) FEBS Lett. 285, 149-151) using NMR techniques. The ionic strength dependence of the observed rate constant for reduction by the methyl viologen radical cation indicates that ionic strength substantially alters the structure and/or the heme reduction potentials of the cytochrome. This result is confirmed by reduction with a neutral flavin species (5-deazariboflavin semiquinone) in which the reactivity of the highest potential heme decreases and the reactivity of the lowest potential heme increases at high (500 mM) ionic strength, and by the sensitivity of heme methyl resonances to ionic strength as observed by 1H-NMR. These unusual ionic strength-dependent effects may be due to a combination of structural changes in the cytochrome and alterations of the electrostatic fields at elevated ionic strengths.
黄素半醌和还原态甲基紫精对四血红素细胞色素c3（来自普通脱硫弧菌，宫崎F株和希尔德堡株）的还原遵循单相动力学曲线，尽管这四个血红素没有相等的还原电位。通过光滴定逐步还原细胞色素后，可得到各个血红素的还原速率常数。每个速率常数对血红素与还原剂之间还原电位差的依赖性可用外层电子转移理论来描述。因此，细胞色素c3血红素极低的还原电位弥补了血红素极大的溶剂可及性。细胞色素c3四个血红素的电子转移相对速率常数与Park等人（Park, J.-S., Kano, K., Niki, S. and Akutsu, H. (1991) FEBS Lett. 285, 149 - 151）先前使用NMR技术对血红素还原电位的分配一致。甲基紫精自由基阳离子还原所观察到的速率常数对离子强度的依赖性表明，离子强度会显著改变细胞色素的结构和/或血红素还原电位。用中性黄素物种（5 - 脱氮核黄素半醌）进行还原可证实这一结果，在高离子强度（500 mM）下，最高电位血红素的反应性降低，最低电位血红素的反应性增加，并且通过1H - NMR观察到血红素甲基共振对离子强度的敏感性。这些不寻常的离子强度依赖性效应可能是由于细胞色素结构变化和离子强度升高时静电场改变共同作用的结果。
