Matsumoto Yushi, Muneyuki Eiro, Fujita Daisuke, Sakamoto Kimitoshi, Miyoshi Hideto, Yoshida Masasuke, Mogi Tatsushi
Chemical Resources Laboratory, Tokyo Institute of Technology, Nagatsuta 4259, Midori-ku, Yokohama 226-8503.
J Biochem. 2006 Apr;139(4):779-88. doi: 10.1093/jb/mvj087.
Cytochrome bd is a heterodimeric terminal ubiquinol oxidase of Escherichia coli under microaerophilic growth conditions. The oxidase activity shows sigmoidal concentration-dependence with low concentrations of ubiquinols, and a marked substrate inhibition with high concentrations of ubiquinol-2 analogs [Sakamoto, K., Miyoshi, H., Takegami, K., Mogi, T., Anraku, Y., and Iwamura H. (1996) J. Biol. Chem. 271, 29897-29902]. Kinetic analysis of the oxidation of the ubiquinol-2 analogs, where the 2- or 3-methoxy group has been substituted with an azido or ethoxy group, suggested that its peculiar enzyme kinetics can be explained by a modified ping-pong bi-bi mechanism with the formation of inactive binary complex FS in the one-electron reduced oxygenated state and inactive ternary complex (E2S)S(n) on the oxidation of the second quinol molecule. Structure-function studies on the ubiquinol-2 analogs suggested that the 6-diprenyl group and the 3-methoxy group on the quinone ring are involved in the substrate inhibition. We also found that oxidized forms of ubiquinone-2 analogs served as weak noncompetitive inhibitors. These results indicate that the mechanism for the substrate oxidation by cytochrome bd is different from that of the heme-copper terminal quinol oxidase and is tightly coupled to dioxygen reduction chemistry.
细胞色素bd是大肠杆菌在微需氧生长条件下的一种异源二聚体末端泛醇氧化酶。该氧化酶活性在低浓度泛醇时呈S形浓度依赖性，而在高浓度泛醇-2类似物时表现出明显的底物抑制作用[坂本，K.，三好，H.，武上，K.，茂木，T.，安楽，Y.，和岩村，H.（1996）《生物化学杂志》271，29897 - 29902]。对泛醇-2类似物氧化的动力学分析表明，其中2-或3-甲氧基已被叠氮基或乙氧基取代，其独特的酶动力学可以用一种修正的乒乓双底物机制来解释，即在单电子还原的氧化态下形成无活性的二元复合物FS，在第二个喹醇分子氧化时形成无活性的三元复合物（E2S）S(n)。对泛醇-2类似物的结构-功能研究表明，醌环上的6-二异戊烯基和3-甲氧基参与了底物抑制作用。我们还发现泛醌-2类似物的氧化形式作为弱非竞争性抑制剂。这些结果表明，细胞色素bd氧化底物的机制不同于血红素-铜末端喹醇氧化酶，并且与双氧还原化学紧密偶联。
