Mogi Tatsushi
Department of Biomedical Chemistry, Graduate School of Medicine, University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
J Biochem. 2009 Jun;145(6):763-70. doi: 10.1093/jb/mvp033. Epub 2009 Mar 2.
Cytochrome bd is a cyanide-resistant terminal quinol oxidase under micro-aerophilic growth conditions and generates a proton motive force via scalar protolytic reactions. Protons used for dioxygen reduction are taken up from the cytoplasm and delivered to haem d through a proton channel. Electrons are transferred from quinols to haem d through haem b558 and haem b595. All three haems are bound to subunit I but only the axial ligand of haem d remains to be determined. Haems b595 and d form a haem-haem binuclear centre and substitutions of either His19 in helix I (haem b595 ligand) and Glu99 in helix III eliminated or severely reduced both haems. To probe the location of the haem d ligand, we introduced mutations around His19 and Glu99 and examined the cyanide-resistance of the oxidase activity and spectroscopic properties. In contrast to mutations around His19, I98F and L101T reduced the IC50 for cyanide to 0.18 and 0.41 mM, respectively, from 1.4 mM of the wild-type. Blue shifts in the alpha peak of I98F suggest that Ile98 is in the vicinity of the haem d-binding site. Our data are consistent with the proposal that Glu99 serves as a haem d ligand of cytochrome bd.
细胞色素bd是一种在微需氧生长条件下对氰化物有抗性的末端醌氧化酶，通过标量质子解反应产生质子动力。用于氧气还原的质子从细胞质中摄取，并通过质子通道传递到血红素d。电子从醌类通过血红素b558和血红素b595转移到血红素d。所有这三种血红素都与亚基I结合，但只有血红素d的轴向配体有待确定。血红素b595和d形成一个血红素-血红素双核中心，螺旋I中的His19（血红素b595配体）和螺旋III中的Glu99的取代消除或严重降低了这两种血红素。为了探究血红素d配体的位置，我们在His19和Glu99周围引入突变，并检测氧化酶活性的抗氰性和光谱性质。与His19周围的突变不同，I98F和L101T将氰化物的IC50分别从野生型的1.4 mM降低到0.18和0.41 mM。I98F的α峰蓝移表明Ile98在血红素d结合位点附近。我们的数据与Glu99作为细胞色素bd的血红素d配体的提议一致。
