YopT: A Yersinia Virulence Factor

YopT：耶尔森菌毒力因子

• 批准号: 7052060
• 负责人: JACK E DIXON
• 金额: $ 30.15万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-04-15 至 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7052060
• 关键词: X ray crystallography; Yersinia pestis; Yersinia pestis disease; antibiotics; bacteria infection mechanism; bacterial proteins; bioassay; bioterrorism /chemical warfare; chemical models; drug resistance; endopeptidases; enzyme activity; enzyme mechanism; enzyme structure; enzyme substrate complex; genetic strain; host organism interaction; protease inhibitor; protein purification; recombinant proteins; virulence

DESCRIPTION (provided by applicant): Recent acts of bioterrorism have underscored the importance of understanding the molecular mechanisms of bacterial pathogenesis. Yersinia pestis, the causative agent of the plague or the Black Death, has been used previously as a biological weapon. There is also concern that genetic engineering has been used to generate antibiotic resistant strains of Yersinia. This proposal focuses on understanding the biochemistry of the Yersinia virulence factor known as YopT. Our laboratory has recently demonstrated that YopT is a cysteine protease. We propose to develop a highly efficient and sensitive assay for YopT's catalytic activity. We will also produce and purify recombinant YopT, a YopT/SycT complex, and a catalytically inactive YopT complexed with substrate. The purified protein(s) will be used to obtain crystalline protein suitable for structural analyses using X-ray diffraction. We have chosen the R21 venue for funding because, although we have made good progress on determining the function of this virulence protein, we have no preliminary data about its catalytic properties. In addition, even though we have determined the structure of a YopT family member, AvrPphB, it will be essential to have a structure of YopT in order to identify potential inhibitors directed against this protease.

描述（由申请人提供）：生物恐怖主义的最新动作强调了理解细菌发病机理的分子机制的重要性。鼠疫或黑死亡的病原体耶尔西尼亚·佩斯蒂斯（Yersinia Pestis）以前已被用作生物武器。还担心基因工程已被用来产生耶尔森氏菌的抗生素抗性菌株。该提案的重点是理解Yersinia毒力因子的生物化学，称为Yopt。我们的实验室最近证明YOPT是半胱氨酸蛋白酶。我们建议为YOPT的催化活性开发高效和敏感的测定法。我们还将生产并纯化重组YOPT，YOPT/SYCT复合物以及与底物复合的催化性YOPT。纯化的蛋白质将用于获得适合使用X射线衍射进行结构分析的结晶蛋白。我们选择了R21的资金场所，因为尽管我们在确定这种毒力蛋白的功能方面取得了良好的进展，但我们没有有关其催化特性的初步数据。此外，即使我们已经确定了Yopt家族成员AvrpphB的结构，但必须具有Yopt结构，以确定针对该蛋白酶的潜在抑制剂。