CHAPERONES AND THE BIOGENESIS OF MEMBRANE PROTEINS

尽管近年来的许多工作都集中在管腔的作用上 分泌蛋白的折叠和组装中的内质网伴侣，许多 这些蛋白质还具有相当长的胞浆部分。假设是这样的 他们还需要监护人的行动，但这还没有 详细调查过了。ABC蛋白家族的成员都有非常 长的胞浆结构域，包含蛋白质的大部分。它 是公认的dna J同系物与dna K同系物相互作用 (Hsp70)促进底物蛋白折叠。胞质，内质网 在酵母和酵母中发现了定位的dna J同源物(Hsp40s) 哺乳动物。这一建议的假设是YDJ-1(酵母)和HDJ- 1和-2(哺乳动物)与胞浆HSP70合作折叠 胞浆尾部的内质网膜蛋白。在第一个目标中，私人侦探将 利用遗传和生化技术检测YDJ-1和 HSP70和Ste6在酿酒酵母中折叠。在第二个目标中，他将 开发一种无细胞体系来表征反应中间体 CFTR折叠的途径。在第三个目标中，提纯的成分将被 用来确定HSP40/HSP70的分子机制 伴侣对促进可溶性胞质结构域的折叠 膜蛋白。

Although much work in recent years has been focused on the role of lumenal ER chaperones in the folding and assembly of secretory proteins, many of these proteins also possess rather long cytosolic portions. It is assumed that they also require the action of chaperones, but this has not been investigated in any detail. Members of the ABC protein family have very long cytosolic domains that encompass a major portion of the protein. It is well recognized that dnaJ homologues interact with dnaK homologues (hsp70s) to promote the folding of substrate proteins. Cytosolic, ER localized, dnaJ homologues (hsp40s) have been identified in yeast and mammals. It is the hypothesis of this proposal that YDJ-1 (yeast) and HDJ- 1 and -2 (mammalian) collaborate with cytosolic hsp70 to fold the cytosolic tail of ER membrane proteins. In the first aim, the P.I. will use genetic and biochemical techniques to examine the role of YDJ-1 and hsp70 and Ste6 folding in S. cerevisiae. In the second aim, he will develop a cell free system to characterize reaction intermediates in the pathway for CFTR folding. In the third aim, purified components will be employed to determine the molecular mechanism by which hsp40/hsp70 chaperone pairs facilitate the folding of soluble cytosolic domains on membrane proteins.