Role of Subunit III in Cytochrome Oxidase Function

亚基 III 在细胞色素氧化酶功能中的作用

基本信息

批准号：
7171935
负责人：
JONATHAN P HOSLER
金额：
$ 27.91万
依托单位：
UNIVERSITY OF MISSISSIPPI MED CTR
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-06-01 至 2008-12-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Cytochrome c oxidase is one of the key energy-generating proteins that function to pump protons across the inner membrane of mitochondria. The catalytic core of the enzyme is composed of three integral membrane proteins, subunits I, II and III. Subunit III does not play a direct role in electron transfer, but it does affect the function of the proton transfer pathways and the proton pumping mechanism in subunit I. Subunit III also prevents the oxidase from undergoing rapid suicide inactivation during normal catalytic turnover. High resolution structures of cytochrome oxidase show that subunit III is associated with tightly bound phospholipids. Mutagenesis and phospholipase treatment of cytochrome c oxidase of the bacterium Rhodobacter sphaeroides, which is closely related to the mitochondrial oxidase, indicates that these lipids are necessary for binding subunit III to subunit I. This is explained by the oxidase structures which show that four phospholipids are simultaneously coordinated by conserved residues of both subunit I and subunit III. The first aim of this project is to examine the protein-lipid interactions of cytochrome oxidase through the analysis of site-directed mutants designed to interrupt these interactions. These experiments will yield information about how tightly bound phospholipids function to facilitate the binding of membrane proteins. These studies should also reveal what type of inter-subunit interactions between integral membrane proteins are used to transmit structural information. A second aim is to study the process of long-distance proton transfer in cytochrome oxidase, using site-directed mutagenesis of subunit III and subunit I residues. Key questions here are how the protein surface around the entry site of a proton pathway controls the rate of proton transfer and how sensitive is the proton conductive pathway in the interior of the protein to modification of residues that connect the pathway to another subunit. A third aim is to establish what structural changes occur with suicide inactivation in order to understand the mechanism of the inactivation process. A fourth aim is to characterize a copper chaperone that appears to be involved in the insertion of CuB into heme a3-CuB active site of cytochrome oxidase. The mechanism of this assembly process is likely to differ from the assembly of other copper centers in proteins since the active site of cytochrome oxidase is buried within subunit I and within the transmembrane region of the protein.

描述（由申请人提供）：细胞色素C氧化酶是泵送质子跨线粒体内膜的关键产生蛋白之一。酶的催化核心由三个积分膜蛋白，亚基I，II和III组成。亚基III在电子转移中不发挥直接作用，但它确实会影响质子转移途径的功能和亚基I.亚基III中的质子泵送机制，也可以防止氧化酶在正常催化失误期间快速自杀自杀。细胞色素氧化酶的高分辨率结构表明亚基III与紧密结合的磷脂有关。与线粒体氧化酶密切相关的细菌细菌Sphaeroides的细胞色素C氧化酶的诱变和磷脂酶处理表明，这些脂质对于将亚基III与亚基结合到亚基I是必不可少的。由亚基I和亚基III的保守残基同时协调。该项目的第一个目的是通过分析旨在中断这些相互作用的位置定向突变体来检查细胞色素氧化酶的蛋白质脂质相互作用。这些实验将产生有关如何紧密结合的磷脂功能以促进膜蛋白结合的信息。这些研究还应揭示积分膜蛋白之间哪种类型的亚基间相互作用来传输结构信息。第二个目的是使用亚基III和亚基I残基的定位诱变研究细胞色素氧化酶中长距离质子转移的过程。这里的关键问题是质子途径进入位点周围的蛋白质表面如何控制质子转移的速率以及蛋白质内部质子导电途径的敏感性如何，以修饰将途径连接到另一个亚基的残基的修饰。第三个目的是确定自杀失活的结构变化，以了解失活过程的机制。第四个目的是表征铜伴侣，该铜伴侣似乎参与了幼崽插入细胞色素氧化酶的血红素A3-cub活性位点。由于细胞色素氧化酶的活性位点埋在亚基I内，并且在蛋白质的跨膜区域内，因此该组装过程的机制可能与蛋白质其他铜中心的组装有所不同。

项目成果

期刊论文数量（16）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

The roles of Rhodobacter sphaeroides copper chaperones PCu(A)C and Sco (PrrC) in the assembly of the copper centers of the aa(3)-type and the cbb(3)-type cytochrome c oxidases.

球形红杆菌铜伴侣 PCu(A)C 和 Sco (PrrC) 在 aa(3) 型和 cbb(3) 型细胞色素 c 氧化酶铜中心组装中的作用。

DOI：
10.1016/j.bbabio.2012.01.003
发表时间：
2012
期刊：
Biochimica et biophysica acta
影响因子：
0
作者：
Thompson,AudieK;Gray,Jimmy;Liu,Aimin;Hosler,JonathanP
通讯作者：
Hosler,JonathanP

Alternative initial proton acceptors for the D pathway of Rhodobacter sphaeroides cytochrome c oxidase.

球形红细菌细胞色素 c 氧化酶 D 途径的替代初始质子受体。

DOI：
10.1021/bi102002v
发表时间：
2011
期刊：
Biochemistry
影响因子：
2.9
作者：
Varanasi,Lakshman;Hosler,Jonathan
通讯作者：
Hosler,Jonathan

Altering conserved lipid binding sites in cytochrome c oxidase of Rhodobacter sphaeroides perturbs the interaction between subunits I and III and promotes suicide inactivation of the enzyme.

改变球形红杆菌细胞色素 c 氧化酶中保守的脂质结合位点会扰乱亚基 I 和 III 之间的相互作用，并促进该酶的自杀失活。

DOI：
10.1021/bi061390q
发表时间：
2006
期刊：
Biochemistry
影响因子：
2.9
作者：
Varanasi,Lakshman;Mills,Denise;Murphree,Anna;Gray,Jimmy;Purser,Chris;Baker,Rodney;Hosler,Jonathan
通讯作者：
Hosler,Jonathan

Slow proton transfer through the pathways for pumped protons in cytochrome c oxidase induces suicide inactivation of the enzyme.

通过细胞色素c氧化酶中泵送质子的途径缓慢的质子转移会导致酶的自杀性失活。