Kobayashi Kazuo, Tagawa Seiichi, Mogi Tatsushi
Institute of Scientific and Industrial Research, Osaka University, Mihogaoka, Ibaraki, Osaka, Japan.
J Biochem. 2009 May;145(5):685-91. doi: 10.1093/jb/mvp026. Epub 2009 Feb 13.
The Escherichia coli cytochrome bo is a heme-copper terminal ubiquinol oxidase, and functions as a redox-driven proton pump. We applied pulse radiolysis technique for studying the one-electron reduction processes in the Cu(B)-deficient mutant, His333Ala. We found that the Cu(B) deficiency suppressed the heme b-to-heme o electron transfer two order of the magnitude (4.0 x 10(2) s(-1)), as found for the wild-type enzyme in the presence of 1 mM KCN (3.0 x 10(2) s(-1)). Potentiometric analysis of the His333Ala mutant revealed the 40 mV decrease in the E(m) value for low-spin heme b and the 160 mV increase in the E(m) value of high-spin heme o. Our results indicate that Cu(B) not only serves as one-electron donor to the bound dioxygen upon the O-O bond cleavage, but also facilitates dioxygen reduction at the heme-copper binuclear centre by modulating the E(m) value of heme o through magnetic interactions. And the absence of a putative OH(-) bound to Cu(B) seems not to affect the uptake of the first chemical proton via K-channel in the His333Ala mutant.
大肠杆菌细胞色素bo是一种血红素-铜末端泛醇氧化酶，作为氧化还原驱动的质子泵发挥作用。我们应用脉冲辐解技术研究了铜（B）缺陷型突变体His333Ala中的单电子还原过程。我们发现，铜（B）缺陷使血红素b到血红素o的电子转移受到抑制，幅度达两个数量级（4.0×10² s⁻¹），这与在1 mM KCN存在下野生型酶的情况（3.0×10² s⁻¹）相同。对His333Ala突变体的电位分析表明，低自旋血红素b的E(m)值下降了40 mV，高自旋血红素o的E(m)值增加了160 mV。我们的结果表明，铜（B）不仅在O - O键断裂时作为结合双氧的单电子供体，还通过磁相互作用调节血红素o的E(m)值，促进血红素-铜双核中心的双氧还原。并且在His333Ala突变体中，推测与铜（B）结合的OH⁻的缺失似乎不影响通过钾通道摄取第一个化学质子。
