Molecular genetics of Intracellular Protein Transport

描述（由申请人提供）：内质网（ER）是膜和分泌蛋白修饰、折叠和组装的隔室。大多数细胞外蛋白的折叠过程的一部分包括二硫键的形成，二硫键可以增加折叠多肽的稳定性，并可以连接蛋白质复合物的亚基。二硫键的形成需要在ER腔中产生二硫键的酶以及将这些键转移到底物蛋白的途径。凯泽博士的研究小组提出了一个结合生物化学，结构和遗传实验的酵母S。酿酒酵母，使基本的洞察机制的二硫键形成活细胞。Kaiser博士实验室以前的工作已经描绘了ER中蛋白质二硫键形成的核心途径，其中管腔氧化酶Ero1p（或Erv2p）将二硫键转移到蛋白质二硫键异构酶（PDI），后者又将其二硫键转移到底物蛋白质。Ero1p和Erv2p的结构研究表明，尽管这些蛋白质在序列上不相似，但它们具有关键的结构特征，从而可以深入了解二硫键的产生和从一种蛋白质转移到另一种蛋白质的机制。在本申请中，Kaiser博士建议确定二硫键如何选择性地从Ero1p转移到PDI。额外的实验旨在了解ER氧化酶的功能如何整合到细胞的氧化还原生物化学中。酵母中二硫键形成的研究将扩展到了解其他生物（包括G. lamblia和C.优雅 In S.通过研究酿酒酵母，将有可能应用发育良好的遗传生物的全部力量来揭示ER中负责蛋白质折叠的基因和蛋白质。对S.酿酒酵母的发现将使人们有可能了解哺乳动物细胞中的平行过程，开辟了诊断哺乳动物细胞内质网功能障碍性折叠的途径，并提供了控制胞外蛋白组装和分泌的新机会。

DESCRIPTION (provided by applicant): The endoplasmic reticulum (ER) is the compartment where membrane and secretory proteins are modified, folded, and assembled. Part of the folding process for most extracellular proteins includes formation of disulfide bonds which can add stability to folded polypeptides and can link subunits of protein complexes. The formation of disulfide bonds requires both enzymes for the generation of disulfide bonds in the lumen of the ER as well as a pathway for the transfer of these bonds to substrate proteins. Dr. Kaiser's research group proposes a combination of biochemical, structural, and genetic experiments in the yeast S. cerevisiae to give fundamental insight into the mechanisms of disulfide bond formation in living cells. Previous work in Dr. Kaiser's laboratory has delineated the core pathway in the ER for protein disulfide bond formation in which a luminal oxidase Ero1p (or Erv2p) transfers a disulfide bond to protein disulfide isomerase (PDI) which in turn transfers its disulfide bond to a substrate protein. Structural studies of Ero1p and Erv2p reveal that although these proteins are not similar in sequence, they nevertheless share key structural features giving insight into the mechanisms by which disulfide bonds are generated and transferred from one protein to another. In this application, Dr. Kaiser proposes to determine how disulfide bonds are selectively transferred from Ero1p to PDI. Additional experiments are designed to understand how the functions of ER oxidases are integrated into the redox biochemistry of the cell. Studies of disulfide bond formation in yeast will be extended to understand interesting disulfide bond forming processes in other organisms including G. lamblia and C. elegans. In S. cerevisiae it will be possible to apply the full power of a well developed genetic organism to uncover the genes and proteins responsible protein folding in the ER. A detailed understanding of these pathways in S. cerevisiae will make it possible to understand parallel processes in mammalian cells, opening the way to diagnose dysfunctional folding in the ER of mammalian cells and providing new opportunities to control the assembly and secret on of extracellular proteins.