The Role of ATP Hydrolysis in HSV-1 Terminase Activity

ATP 水解在 HSV-1 终止酶活性中的作用

• 批准号: 6801109
• 负责人: Carol Lynn Duffy
• 金额: $ 5.05万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-01 至 2005-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6801109
• 关键词: Escherichia coli; adenosinetriphosphatase; artificial chromosomes; bioenergetics; cell line; chemical kinetics; endodeoxyribonuclease; enzyme activity; enzyme structure; functional /structural genomics; herpes simplex virus 1; hydrolysis; postdoctoral investigator; protein protein interaction; protein purification; protein structure function; recombinant virus; virus DNA; virus assembly; virus protein

DESCRIPTION (provided by applicant): The goal of this project is a thorough analysis of the ATPase activity associated with the herpes simplex virus type 1 packaging enzyme, or terminase. The process of DNA packaging is both conserved among members of the Herpesviridae and distinct from cellular processes. Thus studies on the HSV-1 terminase will provide knowledge important for the generation of anti-viral agents for herpesviruses associated with a number of life-threatening diseases. The hydrolysis of ATP is required for herpesvirus DNA packaging. The UL15 and UL28 proteins have been proposed to comprise the HSV-1 terminase and, therefore, to possess the many activities required for DNA packaging including ATPase activity. The specific aims of this proposal utilize biochemical and genetic approaches to identify and analyze the ATPase center of the HSV-1 terminase. Purified UL15 and UL28 proteins will be tested for ATPase activity and the kinetics of ATP hydrolysis will be determined. ATP-interacting residues will be identified through the sequencing of protease-derived peptides from 8-N3-[alpha-32p]ATP-photolabeled protein. Proteins containing amino acid changes in ATP-interacting residues will be generated, purified, and studied to confirm the importance of those residues to ATP hydrolysis. Finally, HSV-1 mutants carrying the above mutations will be generated and examined in vivo to identify specific DNA packaging activities that require the newly discovered ATPase center. These studies will greatly increase the understanding of the process of DNA packaging by all herpesviruses and will extend our knowledge of ATP-driven molecular motors.

描述（由申请人提供）：这个项目的目标是彻底分析与单纯疱疹病毒1型包装酶或终止酶相关的atp酶活性。DNA包装过程在疱疹病毒科成员之间是保守的，并且与细胞过程不同。因此，对HSV-1末端酶的研究将为与许多危及生命的疾病相关的疱疹病毒的抗病毒药物的产生提供重要的知识。ATP的水解是疱疹病毒DNA包装所必需的。已经提出UL15和UL28蛋白包含HSV-1端酶，因此具有DNA包装所需的许多活性，包括atp酶活性。本提案的具体目的是利用生物化学和遗传学方法来鉴定和分析HSV-1末端酶的atp酶中心。纯化的UL15和UL28蛋白将测试ATP酶活性，并确定ATP水解动力学。atp相互作用残基将通过8-N3-[α -32p] atp光标记蛋白的蛋白酶衍生肽测序来鉴定。在ATP相互作用残基中含有氨基酸变化的蛋白质将被生成、纯化和研究，以确认这些残基对ATP水解的重要性。最后，将产生携带上述突变的HSV-1突变体，并在体内进行检测，以确定需要新发现的atp酶中心的特定DNA包装活性。这些研究将大大增加对所有疱疹病毒DNA包装过程的理解，并将扩展我们对atp驱动的分子马达的认识。