Determinants of Membrane Protein Topology

描述（由申请人提供）： E.大肠杆菌乳糖通透酶是一种十二跨膜结构域蛋白，其显示依赖于膜的脂质组成的可变拓扑结构。拓扑结构是动态的，随着脂质环境的变化而改变。跨膜螺旋数VII似乎是这些转变的一个铰链点，可能是因为它的相对亲水性，和noncanonical extrammembrane loops之前it.In这个建议中，我们概述了工作，将确定特定的残基乳糖通透酶，有助于拓扑灵活性。将通过定点诱变和遗传筛选鉴定导致脂质非依赖性拓扑结构或新脂质依赖性拓扑结构特征的氨基酸变化。将评估脂质与这些新鉴定的蛋白质突变体的相互作用。从这些研究中，我们希望确定重要的脂质相关因素，在确定初始拓扑结构和拓扑结构的灵活性。乳糖通透酶折叠的一般原理也适用于大肠杆菌和大肠杆菌中的其他膜蛋白。大肠杆菌和真核生物，从而有助于寻找诸如阿尔茨海默氏病和囊性纤维化的膜蛋白紊乱的有效疗法。

DESCRIPTION (provided by applicant): E. coli lactose permease is a twelve-transmembrane domain protein that displays variable topology dependent on the lipid composition of the membrane. The topology is dynamic, changing posttranslationally and post-insertionally with changes in the lipid environment. Transmembrane helix number VII seems to be a hinge point for these transitions, likely because of its relatively hydrophilic nature, and the noncanonical extramembrane loops that precede it. In this proposal, we outline work that will identify specific residues in lactose permease that contribute to topological flexibility. Amino acid changes that lead to either lipid-independent topologies or new lipid-dependent topology profiles will be identified by both site directed mutagenesis and genetic screens. Lipid interactions with these newly identified protein mutants will be assessed. From these studies, we hope to identify lipid-related factors important in determining initial topology and topological flexibility. The general principles governing the folding of lactose permease will apply to other membrane proteins in both E. coli and eukaryotes, and thereby aid in the search for effective therapies of such membrane protein disorders as Alzheimer's Disease and cystic fibrosis.