STRUCTURAL AND FUNCTIONAL STUDIES OF HSP40

HSP40 的结构和功能研究

基本信息

批准号：
8000171
负责人：
BINGDONG SHA
金额：
$ 8.5万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-01-01 至 2010-07-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): The long-term goat of this proposal is to carry out X-ray crystallographic studies on Hsp40 to uncover the basic mechanisms by which Hsp40 interacts with non-native polypeptides and cooperates with Hsp70 to perform the molecular chaperone activity. We have proposed an "anchoring and docking" model to illustrate the mechanisms for Hsp40 to interact with HspT0. To support this hypothesis, we intend to determine the crystal structure of the yeast type II Hsp40 Sis1 complexed with yeast Hsp70 Ssa1. We have constituted and crystallized the protein complex of Sis1 peptide-binding fragment and the Ssal C-terminal domain. Once solved, the complex crystal structure of Sis1 and Ssa1 will inform us how Hsp40 interacts with Hsp70 to carry out the non-native polypeptide delivery and assists Hsp in protein folding. The crystal structure of yeast type I Hsp40 Ydjl, complexed with the peptide substrate GWLYEIS, indicates that the pocket on the molecular surface of Ydjl may play important roles in peptide substrate binding. This pocket may exhibit significant plasticity to accommodate various sizes of hydrophobic side chains. We propose to determine the crystal structures of Ydjl peptide-binding fragment, complexed with peptide substrates GWWYEIS, and GWAYEIS. The crystal structures of Ydj1 and various peptide substrates may provide the structural basis of how Hsp40 adjusts itself to accommodate different peptide substrates. To reveal the mechanism by which Hsp40 primes the non-native polypeptide for the subsequent Hsp70 binding, we have designed a non-native polypeptide, Y1, by connecting two Hsp40 Ydj1 peptide substrates GWLYEIS with a flexible linker. The recombinant polypeptide Y1 showed a reasonable binding affinity to Hsp40 Ydjl. We intend to crystallize and determine the structure of the Ydjl complexed with the "non-native polypeptide", Y1, to illustrate how Hsp40 may affect the conformation of the non-native polypeptide. Finally, we will conduct structure-based mutagenesis studies to test our proposed models for the mechanisms of Hsp40 chaperone actions.

描述（申请人提供）：本提案的长期目标是对Hsp40进行X射线晶体学研究，揭示Hsp40与非天然多肽相互作用并与Hsp70配合发挥分子伴侣活性的基本机制。我们提出了“锚定和对接”模型来说明 Hsp40 与 HspT0 相互作用的机制。为了支持这一假设，我们打算确定与酵母 Hsp70 Ssa1 复合的酵母 II 型 Hsp40 Sis1 的晶体结构。我们构建并结晶了 Sis1 肽结合片段和 Ssal C 端结构域的蛋白质复合物。一旦解决，Sis1和Ssa1的复杂晶体结构将告诉我们Hsp40如何与Hsp70相互作用以进行非天然多肽递送并协助Hsp进行蛋白质折叠。酵母 I 型 Hsp40 Ydjl 与肽底物 GWLYEIS 复合的晶体结构表明，Ydjl 分子表面的口袋可能在肽底物结合中发挥重要作用。该口袋可以表现出显着的可塑性以适应各种尺寸的疏水性侧链。我们建议确定与肽底物 GWWYEIS 和 GWAYEIS 复合的 Ydjl 肽结合片段的晶体结构。 Ydj1和各种肽底物的晶体结构可能为Hsp40如何调整自身以适应不同的肽底物提供结构基础。为了揭示 Hsp40 引发非天然多肽以进行随后的 Hsp70 结合的机制，我们通过使用柔性接头连接两个 Hsp40 Ydj1 肽底物 GWLYEIS 设计了非天然多肽 Y1。重组多肽Y1对Hsp40 Ydjl表现出合理的结合亲和力。我们打算结晶并确定与“非天然多肽”Y1复合的Ydjl的结构，以说明Hsp40如何影响非天然多肽的构象。最后，我们将进行基于结构的诱变研究，以测试我们提出的 Hsp40 伴侣作用机制模型。

项目成果

期刊论文数量（22）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

The crystal structure of the yeast Hsp40 Ydj1 complexed with its peptide substrate.

酵母 Hsp40 Ydj1 与其肽底物复合的晶体结构。

DOI：
发表时间：
2003-12
期刊：
影响因子：
0
作者：
Li, Jingzhi;Qian, Xinguo;Sha, Bingdong
通讯作者：
Sha, Bingdong

The structural plasticity of Tom71 for mitochondrial precursor translocations.

Tom71 对线粒体前体易位的结构可塑性。

DOI：
10.1107/s1744309110025522
发表时间：
2010-09-01
期刊：
Acta crystallographica. Section F, Structural biology and crystallization communications
影响因子：
0
作者：
Jingzhi Li;W. Cui;B. Sha
通讯作者：
B. Sha

Preliminary X-ray crystallographic studies of yeast Hsp40 Ydj1 complexed with its peptide substrate.

酵母 Hsp40 Ydj1 与其肽底物复合的初步 X 射线晶体学研究。