STRUCTURE AND FUNCTION OF BACTERIORHODOPSIN

细菌视紫红质是一种分子量为27，000道尔顿的膜蛋白， 盐生盐杆菌的紫色膜被定义为高 膜的分辨电子衍射，以及具有重离子的膜的分辨电子衍射， 共价连接的金属基团。 基于我们以前的研究，我们可以 现在按顺序确定这些标记的附着位点， 结构 因此，主要目标是确定顺序， 其中七个跨膜序列出现在膜中的方向。 结构 我们以前已经确定了电子的相位 衍射振幅达到实验上的3.7埃（即，从图像）， 并在-120 ℃下记录到2.65埃的衍射，这对 差分映射 我们定义了一个扩展分辨率7 x10埃 三维结构，显示有助于建立模型的附加功能 蛋白质结构：在确定了视网膜附着位点后， 序列，我们将确定参与质子转移机制的侧链 使用位点特异性诱变的蛋白质泵送，并使用突变体， 研究在H.加州大学旧金山分校的博耶。 的 具体场地的变化将以结构为基础， 将评估变更和功能差异，以确定 光能转换，插入膜，膜蛋白 稳定性，脯氨酸的作用，螺旋之间的肽的作用， 水相，并与最终目标的理解机制， 一种典型的跨膜蛋白介导的现象。 增加 与眼睛中的视紫红质的相似性正在显现。 总体目标是 了解膜介导的现象，这种蛋白质是集中在作为 一个可能比其他人更快产生基本洞察力的原型。

The structure of bacteriorhodopsin, a 27,000 dalton membrane protein from purple membrane of Halobacterium halobium is to be defined by high resolution electron diffraction of membranes, and of membranes with heavy metal groups covalently attached. Building on our previous studies, we can now determine the site of attachment of such labels in sequence and structure. Thus a primary objective is to determine the order and orientation in which the seven transmembrane sequences occurs in the structure. We have previously determined the phases of electron diffraction amplitudes to 3.7 Angstroms experimentally (i.e., from images), and recorded diffraction to 2.65 Angstroms at -120 C and this is crucial to difference mapping. We have defined an extended resolution 7x10 Angstrom 3-D structure that shows additional features helpful in building a model of the protein structure: Having defined the retinal attachment site in sequence, we will identify side chains involved in the mechanism of proton pumping using site-specific mutagenesized proteins, and using mutants under study in a closely complementary proposal of H. Boyer at UCSF. The site-specific changes will be based on structure, and both structural change and functional variance will be assessed to define a mechanism of light-energy conversion, insertion into the membrane, membrane protein stability, the role of prolines, the role of peptides between helices in the aqueous phase, and with the end goal of understanding the mechanisms in an archetype of transmembrane protein-mediated phenomena. Increasing similarities to rhodopsin in the eye is emerging. An overall goal is to understand membrane mediated phenomena, and this protein is focussed on as an archetype likely to yield much fundamental insight sooner than others.