MOLECULAR VIROLOGY OF THE FELINE IMMUNODEFICIENCY VIRUS PROTEASE

猫免疫缺陷病毒蛋白酶的分子病毒学

• 批准号: 6340975
• 负责人: John H Elder
• 金额: $ 11.38万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-01 至 2001-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6340975
• 关键词: SDS polyacrylamide gel electrophoresis; antiviral agents; chemical cleavage; drug screening /evaluation; endopeptidases; enzyme activity; enzyme substrate; feline immunodeficiency virus; genetic manipulation; high performance liquid chromatography; mass spectrometry; molecular biology; molecular cloning; molecular site; mutant; site directed mutagenesis; synthetic peptide; tissue /cell culture; virus protein

The purpose of the proposed research is to utilize the feline lentivirus, FIV, as a system for testing strategies aimed at the rational design of antivirals efficacious against lentivirus infections. The similarities between FIV and HIV in life cycle, genomic make-up, and etiology of disease resulting from infection make the cat a particularly relevant model. In addition, the relative economy, safety, and manipulability of the feline system make it much more amenable to detailed testing than any of the primate systems. Studies will center around the characterization of FIV protease (PR) and the sites within FIV that are cleaved by this critical enzyme. Both eucaryotic and procaryotic expression clones will be prepared in order to 1) determine PR cleavage sites; 2) establish a hierarchy of cleavage; 3) assess cleavage rates of the natural and synthetic PRs against native substrates; and 4) monitor the influence of drugs arising from Projects 1-4 on Pr cleavage of natural substrates. The data generated will be used for synthetic substrate preparation in Project 4; drug design by projects 1, 2 and 3; for comparison of cleavage specificities with HIV PR; and to aid in the establishment of consensus rules for retroviral PR cleavage. This project will also be responsible for the preparation of site-directed mutants of PR, based on predicted interactions between enzyme and drug, established in Projects 1-4. We will also carry out all virological aspects of the program, which will involve the in vitro testing of selected drugs for both toxicity and antiviral efficacy. The results of these studies will fulfill the anchor phase of the developmental loop, providing support and feedback for design strategies arising from the overall program.

这项拟议的研究的目的是利用猫慢病毒， FIV，作为一种测试策略的系统，旨在合理设计 抗病毒药物对慢病毒感染有效。相似之处 FIV和HIV在生命周期、基因组组成和疾病病因学中的关系 感染导致的疾病使猫成为一个特别相关的模型。在……里面 此外，猫科动物的相对经济性、安全性和可操作性 系统使其更易于进行详细的测试 灵长类系统。研究将围绕FIV的特征展开 蛋白水解酶(PR)和FIV内被这个关键的 酵素。将同时准备真核和原核表达克隆 为了1)确定PR裂解位点；2)建立 卵裂；3)评估天然和合成PR对 天然底物；以及4)监测药物产生的影响 项目1-4关于天然底物的PR裂解。生成的数据将 用于项目4中的合成底物制备；药物设计由 项目1、2和3；比较与艾滋病毒PR的切割特异性； 并协助建立关于逆转录病毒公关的共识规则 乳沟。该项目还将负责筹备 PR的定点突变，基于预测的酶之间的相互作用 和药物，在项目1-4中建立。我们还将执行所有 该计划的病毒学方面，这将涉及体外测试 所选药物的毒性和抗病毒疗效。结果是 这些研究将完成发育环的锚定阶段， 为设计策略提供支持和反馈 总体规划。