UNIQUE AMINO DOMAINS AND AMP DEAMINASE ISOFORM DIVERSITY

独特的氨基结构域和 AMP 脱氨酶异构体多样性

• 批准号: 2760267
• 负责人: RICHARD L SABINA
• 金额: $ 19.39万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-01 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2760267
• 关键词: AMP deaminase; X ray crystallography; actins; active sites; adenylate kinase; affinity labeling; enzyme deficiency; enzyme mechanism; enzyme structure; gene expression; hybrid enzyme; isozymes; muscle metabolism; myosins; phosphatidylinositols; phosphorylation; protein binding; protein isoforms; protein localization; protein protein interaction; protein sequence; protein structure function; recombinant proteins

AMP deaminase (AMPD; EC 3.5.4.6) is a diverse and highly regulated enzyme located at a branchpoint in the adenylate catabolic pathway. AMPD plays a key role during metabolic imbalances of energy supply and demand homeostasis by competing with 5' nucleotidase for available AMP. This role is underscored in striated muscle where inheritance of a prolonged survival in congestive hart failure. These opposing clinical consequences reflect a functional difference in adenylate catabolism in various striated muscles due, in part, to distinct regulatory features of AMPD isoforms. Emerging data show that divergent N-terminal domains in AMPD polypeptides alter physical and functional properties of the enzyme. The long-term objective of this project is to gain a comprehensive understanding of AMPD regulation in striated muscle with an emphasis on the functional significance for divergent N-terminal domains. The continued pursuit of this goal will be accomplished by expressing, isolating an characterizing wild type and genetically modified human AMPD recombinant enzymes that will be used to 1) define the structural bases and functional effects of phosphorylation and phosphoinositides on the catalytic and actomyosin binding properties of human AMPD recombinant isoforms. These analyses will also sere to assess the relationship between these two opposing regulators of AMPD catalytic activity, 2) determine the combined effects of a cassette-type alternative splicing event and a P43L substitution on the human AMPD1 recombinant enzyme. This latter information is central to hypotheses designed to explain clinical outcomes resulting from inheritance of the prevalent AMPD1 mutant allele, and 3) attempt to solve the crystal structure of the human AMPD1 recombinant enzyme. Detailed structural knowledge of AMPD1 will facilitate our understanding of the complex regulation of this enzyme. These combined efforts should provide critical information that may help explain clinical outcomes associated with a prevalent AMPD1 mutant allele.

AMP脱氨酶（AMPD; EC 3.5.4.6）是位于腺苷酸分解代谢途径中的分支点的多样且高度调节的酶。AMPD通过与5'核苷酸酶竞争有效AMP，在能量供需平衡的代谢失衡中起关键作用。这一作用在横纹肌中得到强调，因为横纹肌遗传了充血性哈特衰竭患者的长期存活。这些相反的临床结果反映了腺苷酸催化剂在各种横纹肌中的功能差异，部分原因是AMPD亚型的不同调节特征。新兴的数据表明，不同的N-末端结构域在AMPD多肽改变酶的物理和功能特性。本项目的长期目标是全面了解横纹肌中AMPD的调节，重点是不同N端结构域的功能意义。将通过表达、分离表征野生型和遗传修饰的人AMPD重组酶来实现对该目标的持续追求，所述重组酶将用于1）确定磷酸化和磷酸肌醇对人AMPD重组同种型的催化和肌动球蛋白结合性质的结构基础和功能效应。这些分析还将用于评估AMPD催化活性的这两种相反调节剂之间的关系，2）确定盒式可变剪接事件和P43 L置换对人AMPD 1重组酶的组合效应。后一种信息是设计用于解释由流行AMPD 1突变等位基因遗传引起的临床结果的假设的核心，以及3）试图解决人AMPD 1重组酶的晶体结构。AMPD 1的详细结构知识将有助于我们理解这种酶的复杂调控。这些联合努力应该提供关键信息，可能有助于解释与流行的AMPD 1突变等位基因相关的临床结果。