REGULATION AND FUNCTION OF THE YEAST HEAT SHOCK RESPONSE

蛋白质必须正确地折叠成确定的三维结构才能正常发挥功能。 包括克雅氏病和囊性纤维化在内的几种人类疾病可以被认为是蛋白质 折叠疾病，因为蛋白质不能在其适当的细胞区室中实现活性构象。 在拥挤的细胞环境中，新翻译的蛋白质易于聚集和非生产性 由于暴露于疏水残基的水性环境而过早折叠， 通常隐藏在折叠的蛋白质中。分子伴侣是蛋白质的极好候选物， 由于它们的瞬时性， 与疏水多肽片段的相互作用。在体内和体外的各种系统中收集的数据， 指示从蛋白质在核糖体上是新生链的时间到它们在核糖体上是新生链的时间的蛋白质折叠的途径。 或者适当折叠或者降解。分子伴侣在这些过程中发挥重要作用。 本建议中描述的实验的重点是确定Hsp70（热休克）的功能 蛋白70K）和Hsp40作为分子伴侣，使用芽殖酵母酿酒酵母（Saccharomycescerevisiae 作为一个模型系统。更具体地说，重点是在早期事件的分子伴侣的作用， 核糖体上的蛋白质折叠。S.酿酒酵母含有两类丰富的，功能上 不同的Hsp70（SSA和SSB蛋白）和三种Hsp40（Sisl、Zuol和Ydjl）。在这些监护人中， Sisl和Zuol与翻译核糖体有关，表明在蛋白质生命早期的作用。使用 结合遗传学和生物化学的方法，我们将分析Ssb，Zuol和Sisl在 翻译和蛋白质折叠的早期步骤。 已知Hsp70和Hsp40作为分子伴侣协同起作用。然而，功能 他们之间的关系还不太清楚。我们的目标是了解 与核糖体相关的两个Hsp40（Sisl和Zuol）和一个Hsp70（Ssb）。更 具体地说，我们将通过遗传和生化实验来确定这两种DnaJ在功能上是否 并与Ssb进行物理交互。此外，我们将确定一个新生链的相互作用的顺序 与核糖体相关的伴侣蛋白。

Proteins must fold correctly into a defined three-dimensional structure in order to properly function. Several human diseases, includingCreutzfeldt-Jacob disease and cystic fibrosis, can be considered protein folding diseases because proteins fail to achieve active conformations in their proper cellular compartment. Within the crowded cellular milieu, newly translated proteins are prone to aggregation and nonproductive premature folding because of the exposure to the aqueous environment of hydrophobic residues which are normally sequestered within the folded protein. Molecular chaperones are excellent candidates for proteins which bind to and protect nascent chains from problematic interactions because of the nature of theirtransient interactions with hydrophobic polypeptide segments. Data collected in a variety of systems, in vivo and in vitro, indicate a pathway(s) of protein folding from the time proteins are nascent chains on ribosomes until they are either properly folded or degraded. Molecular chaperones play important roles in these processes. The focus of experiments described in this proposal is to determine the function of Hsp70 (heat shock protein 70K) and Hsp40 as molecular chaperones chaperone using the budding yeast Saccharomyces cerevisiae as a model system. More specifically, the focus is on the role of molecular chaperones during the early events of protein folding on the ribosome. The cytosol of S. cerevisiae contains two classes of abundant, functionally distinct Hsp70s (SSA and SSB proteins) and three Hsp40s (Sisl, Zuol and Ydjl). Of these chaperones, Ssb, Sisl and Zuol associate with translating ribosomes, suggesting a role early in the life of a protein. Using a combination of genetic and biochemical approaches we will analyze the function of Ssb, Zuol and Sisl in translation and early steps of protein folding. Hsp70s and Hsp40s are known to function cooperatively as chaperones. However, the functional relationship between them is not well understood. Our goal is to understand the functional relationship between the two Hsp40s (Sisl and Zuol) and one Hsp70 (Ssb) which are associated with the ribosome. More specifically, we will ascertain through genetic and biochemical experiments whether these two DnaJs functionally and physically interact with Ssb. In addition, we will determine the sequence of interactions of a nascent chain with the ribosome-associated chaperones.