Structure-Function Analysis of Enzymes

酶的结构功能分析

• 批准号: 6749035
• 负责人: Hazel M. Holden
• 金额: $ 29.1万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-06-01 至 2006-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6749035
• 关键词: X ray crystallography; acetyl coA carboxylase; ammonia; aspartate; bacterial proteins; binding sites; biotin; carbamoylphosphate synthase; cofactor; computer simulation; conformation; crystallization; enzyme activity; enzyme complex; enzyme mechanism; enzyme structure; enzyme substrate complex; hydroxymethyltransferases; intermolecular interaction; ligase; molecular dynamics; protein purification; site directed mutagenesis; structural biology

DESCRIPTION: (Applicant's Description) The concept of substrate channeling was originally put forth to explain the manner in which reactive intermediates are transferred from one protein to another in a metabolic pathway or shuttled from one active site to another within a single enzyme. Many have regarded the phenomenon as a critical method for the regulation of metabolic pathways within the living cell. The goal of this proposal is to explore, on a structural basis, the mechanisms of actions of the following enzymatic systems known to employ substrate channeling: carbamoyl phosphate synthetase, asparagine synthetase, and acetyl CoA carboxylase. The first two enzymes utilize substrate channeling between active sites within a single polypeptide chain while in the latter protein, the event occurs between active sites positioned on different polypeptide chains. These systems were selected for study not only because they use substrate channeling in their reaction mechanisms, but also because of their biological importance. Carbamoyl phosphate synthetase is involved in the de novo synthesis of pyrimidine nucleotides which are critical for cell proliferation. The enzyme also plays a key role in both arginine biosynthesis and the urea cycle in most terrestrial vertebrates. Asparagine synthetase catalyzes the ATP-dependent conversion of aspartic acid to asparagine and has been used in chemotherapeutic protocols for treating acute lymphoblastic leukemia. Finally, acetyl CoA carboxylase catalyzes the first committed and absolutely critical step in fatty acid synthesis. For these investigations, a combination of site-directed mutagenesis experiments and x-ray crystallographic analyses will be employed in order to more fully characterize the three-dimensional architectures of these proteins, their active site geometries, catalytic mechanismd, and the relationships of these parameters to substrate channeling.

描述：（申请人的描述）底物沟道的概念是 最初提出是为了解释反应中间体的反应方式 在代谢途径中从一种蛋白质转移到另一种蛋白质或从 单个酶内的一个活性位点到另一个活性位点。很多人都把 现象作为调节体内代谢途径的关键方法 活细胞。该提案的目标是探索结构性的 基础上，已知以下酶系统的作用机制 采用底物通道：氨基甲酰磷酸合成酶、天冬酰胺 合成酶和乙酰辅酶A羧化酶。前两种酶利用底物 单个多肽链内活性位点之间的通道 后一种蛋白质，该事件发生在位于不同位置的活性位点之间 多肽链。选择这些系统进行研究不仅是因为它们 在其反应机制中使用底物通道，但也因为 它们的生物学重要性。氨基甲酰磷酸合成酶参与 从头合成对细胞至关重要的嘧啶核苷酸 增殖。该酶在精氨酸生物合成中也发挥着关键作用 以及大多数陆生脊椎动物的尿素循环。天冬酰胺合成酶 催化天冬氨酸向天冬酰胺的 ATP 依赖性转化，并具有 已用于治疗急性淋巴细胞白血病的化疗方案 白血病。最后，乙酰辅酶A羧化酶催化第一个承诺和 脂肪酸合成中绝对关键的一步。对于这些调查， 定点诱变实验和 X 射线晶体学的结合 将进行分析，以便更全面地描述 这些蛋白质的三维结构及其活性位点 几何形状、催化机制以及这些参数与 底物沟道。

项目成果

Long-range allosteric transitions in carbamoyl phosphate synthetase.

氨基甲酰磷酸合成酶中的长程变构转变。

• 发表时间: 2004-09
• 作者: Thoden, James B;Huang, Xinyi;Kim, Jungwook;Raushel, Frank M;Holden, Hazel M
• 通讯作者: Holden, Hazel M

Antibacterial and Anti-biofilm Activity of the Human Breast Milk Glycoprotein Lactoferrin against Group B Streptococcus.

人母乳糖蛋白乳铁蛋白对 B 族链球菌的抗菌和抗生物膜活性。

• 发表时间: 2021
• 作者: Lu, Jacky;Francis, Jamisha D;Guevara, Miriam A;Moore, Rebecca E;Chambers, Schuyler A;Doster, Ryan S;Eastman, Alison J;Rogers, Lisa M;Noble, Kristen N;Manning, Shannon D;Damo, Steven M;Aronoff, David M;Townsend, Steven D;Gaddy, Jennifer A
• 通讯作者: Gaddy, Jennifer A