Deciphering Enzyme Specificity: Amidohydrolase Superfamily

破译酶的特异性：酰胺水解酶超家族

• 批准号: 7743893
• 负责人: Frank M. Raushel
• 金额: $ 30.6万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7743893
• 关键词: Active Sites; Amides; Amidohydrolases; Amino Acid Sequence; Amino Acids; Biochemical; Biological Assay; C-terminal; Chemicals; Code; Collaborations; Complement; Deamination; Decarboxylation; Detection; Development; Dihydroorotase; Docking; Enzymes; Esters; Evolution; Funding; Genes; Genomics; Goals; Homology Modeling; Hydration status; Hydrolysis; Instruction; Laboratories; Lactones; Libraries; Ligand Binding; Metabolic; Metabolic Pathway; Metals; Methodology; Methods; Modeling; Molecular; Mononuclear; New York; Nucleic Acids; Operon; Organophosphates; Peptide Sequence Determination; Phosphoric Triester Hydrolases; Principal Investigator; Proteins; Protocols documentation; Reaction; Research; Resolution; Roentgen Rays; Screening procedure; Solvents; Specificity; Structure; Substrate Specificity; System; Therapeutic Intervention; Urease; Water; adenosine deaminase; combinatorial; enzyme structure; forging; functional group; insight; member; novel; programs; protein structure; small molecule; small molecule libraries; structural genomics

The long-term objective for the research described in this application is directed at a comprehensive understanding of the physical and chemical parameters that relate protein structure and substrate recognition in enzyme-catalyzed reactions. This objective is aimed toward the development of general methodologies and novel protocols for the determination of reaction and substrate specificities for enzymes of unknown function. A critical assessment of the functional annotations forthe more than four million genes that have been sequenced to date suggests that approximately one-third of the encoded proteins have an uncertain, unknown, or /ncorrecf functional assignment. This observation suggests that a significant fraction of the metabolic diversity remains to be properly characterized. Toward this end we will utilize computational docking of high energy intermediates to models of the active sites for enzymes of unknown function to identify the most probable substrates. These efforts will be complemented by the synthesis and screening of chemical libraries and the abstraction of further metabolic information from operon and genomic context. This goal will be pursued by concentrating on the elucidation of the substrate and reaction profiles forthe entire ensemble of enzymes within the amidohydrolase superfamily. The amidohydrolase superfamily is a group of enzymes which has a substantial substrate diversity embedded within active sites that are forged from a (p/a)8-barrel structural fold. Over 6,000 unique protein sequences have been identified as members of the amidohydrolase superfamily. Members of this superfamily have been shown to catalyze the hydrolysis of amides, lactones and organophosphate esters, in addition to decarboxylation, hydration, and isomerization reactions. However, a substantial fraction of the members of this broad superfamily have an ambiguous substrate and reaction specificity that remains to be unraveled. Members of this superfamily of enzymes include dihydroorotase, urease, and adenosine deaminase. The hallmark for this particular superfamily of enzymes is an active site at the C-terminal end of a (p/a)8-barrel structural domain that contains a mononuclear or binuclear metal center that functions predominantly, but not exclusively, to activate solvent water for nucleophilic attack on electrophilic functional groups. The substrate and reaction diversity contained within this enzyme superfamily will provide unique insights into the molecular mechanisms for the evolution and development of novel enzymatic activities from existing structural templates. RELEVANCE (See instructions); The overall objective of this application is directed towards the development of novel and general methods for the elucidation of function for enzymes with unknown substrates. These efforts will unveil new metabolic transformations and identify new targets for therapeutic intervention.

本申请中描述的研究的长期目标是针对一个全面的 了解与蛋白质结构和底物相关的物理和化学参数 在酶催化反应中的识别。这一目标的目的是发展一般 测定酶的反应和底物特异性的方法和新方案 未知的功能。对超过400万个基因的功能注释进行了严格的评估 迄今为止已经测序的结果表明，大约三分之一的编码蛋白质具有 不确定的、未知的或/ncorrect的功能分配。这一观察表明， 的代谢多样性仍然是适当的特点。为此，我们将利用计算 高能中间体与未知功能酶的活性位点模型的对接， 确定最可能的基质。这些努力将辅之以综合和筛选 化学文库和从操纵子和基因组背景中提取进一步的代谢信息。 这一目标将通过集中于阐明底物和反应概况来实现。 酰胺水解酶超家族中的全部酶。酰胺水解酶超家族是一个 一组酶，具有嵌入在活性位点内的大量底物多样性， 从一个（p/a）8桶结构褶皱。超过6，000种独特的蛋白质序列已被确定为 酰胺水解酶超家族。该超家族的成员已被证明可以催化 酰胺、内酯和有机磷酸酯，除脱羧、水合和异构化外 反应.然而，这个广泛的超家族的相当一部分成员具有模糊的 底物和反应特异性仍然有待解开。这个酶超家族的成员 包括二氢乳清酸酶、尿素酶和腺苷脱氨酶。这个特殊的超家族的标志是 酶是在（p/a）8-桶结构域的C-末端的活性位点，所述8-桶结构域含有 单核或双核金属中心，主要但不排他地起活化溶剂的作用 水用于亲电官能团的亲核攻击。底物和反应的多样性 包含在这个酶超家族将提供独特的见解的分子机制， 从现有的结构模板进化和开发新的酶活性。 相关性（参见说明）; 本申请的总体目标是开发新颖且通用的药物组合物。 用未知底物阐明酶功能的方法。这些努力将揭示新的 代谢转化，并确定新的治疗干预目标。