DECIPHERING ENZYME SPECIFICITY: ENOLASE AND RUBISCO SUPERFAMILIES

破译酶的特异性：烯醇化酶和卢比斯科超家族

• 批准号: 7743892
• 负责人: JOHN A GERLT
• 金额: $ 87.42万
• 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/7743892
• 关键词: Acids; Active Sites; Aldehyde-Lyases; Anions; Biology; C-terminal; Catalysis; Databases; Dehydration; Development; Dipeptidases; Dipeptides; Elements; Enzymes; Evolution; Family; Genome; Homologous Gene; Hydro-Lyases; Instruction; Intervention; Isomerase; Ketoses; Ketosis; Link; Location; Mandelate racemase; Metabolic Pathway; Metabolism; Methionine; Molecular; N-terminal; Operon; Oxidoreductase; Oxygenases; Pathway interactions; Phosphotransferases; Proteins; Protons; Reaction; Research; Research Personnel; Ribulose-Bisphosphate Carboxylase; Role; Set protein; Specificity; Structure; Structure-Activity Relationship; Subgroup; Substrate Specificity; Sugar Acids; base; carboxylate; carboxylation; catalyst; chemical reaction; design; dimer; enolase; enolate; epimerase; expectation; functional group; inorganic phosphate; member; novel; novel therapeutic intervention; oxidation; polypeptide; programs; ribulose; small molecule

Mechanistically diverse superfamilies provide the opportunity to understand the structural bases for divergent evolution of enzyme function. In the enolase superfamily, the reactions are initiated by abstraction of the a-proton of a carboxylate anion substrate to yield a Mg^^-stabilized enolate intermediate; the intermediate is directed to product by an appropriately located active site acid. In the RuBisCO superfamily, the reactions are initiated by abstraction of the a-proton of a ketose 1-phosphate substrate to yield a Mg^'^-stabilized enolate intermediate; the potential fate(s) of the intermediate are pooriy understood but may involve tautomerization, dehydration, oxidation, and/or carboxylation. This project describes structure/function aspects of our integrated sequence-structure-computation strategy for predicting the substrates specificities and, therefore, assigning functions of uncharacterized proteins in both superfamilies. The focus is on proteins that are encoded by operons: the enzymes that catalyze successive steps in a metabolic pathway should share conserved elements of substrate specificity, thereby facilitating identification of the functions of all of the enzymes in the pathway and, therefore, new metabolism. The project is organized in three Specific Aims: Specific Aim 1 focuses on divergent members of the muconate lactoniziing enzyme subgroup of the enolase superfamily (Lys acid/base catalysts at the ends of the second and sixth (J-strands of the barrel domain), including 1) dipeptide epimerases that are encoded by operons that also encode homologues of dipeptidases, and 2) two novel subgroups whose members are expected to catalyze "new" reactions. Specific Aim 2 focuses on divergent members of the mandelate racemase subgroup of the enolase superfamily (an acid/base His-Asp dyad at the ends of the seventh and sixth p-strands of the TIM-barrel domain) that are encoded by operons, with these also encoding aldolases, dehydrogenases, mutarotases, and/or kinases. Specific Aim 3 focuses on RuBisCO-like proteins (RLPs) that are encoded by operons that also encode homologues of isomerases, aldolases, transketolases, and other aldose/ketose 5-phosphate utilizing enzymes. RELEVANCE (See instructions); The assignment of functions to the complete set of proteins encoded by genomes is a major problem. However, when this problem is solved, their roles in molecular, cellular, and organismal functions will be known and novel targets for specific small molecule intervention can be identified, thereby providing new approaches for therapeutic design. This Program Project is focused on developing and implementing an integrated sequence-structure-computation strategy for predicting the substrate specificities of uncharacterized proteins discovered in genome projects, thereby facilitating their functional assignment.

机械上多样化的超家族为了解结构基础提供了机会 酶功能的发散演变。在烯醇酶超家族中，反应是通过抽象引发的 羧酸盐阴离子底物的A-proton的含量，得出Mg ^^ - 稳定的中间体；这 中间体由适当位置的活性位点酸引向产品。在鲁比斯科超家族中 反应是通过酮1-磷酸基底物的A-Proton的抽象来引发的，以产生A mg^'^ - 稳定的烯烃中间体；中间体的潜在命运已被理解，但可以 涉及互构化，脱水，氧化和/或羧化。该项目描述 我们集成序列结构策略的结构/功能方面，用于预测 底物特异性，因此分配了两个超家族中未表征的蛋白质的功能。 重点是由操纵子编码的蛋白质：催化连续步骤的酶 代谢途径应具有底物特异性的保守元素，从而促进识别 途径中所有酶的功能，因此是新的新陈代谢。 该项目以三个具体目的组织： 特定目标1的重点是粘液酸酯乳酸化酶亚组的分歧成员 烯醇酶超家族（第二和第六的末端的乳酸/碱催化剂 域），包括1）由操纵子编码的二肽分配酶 二肽酶和2）两个新型亚组，其成员有望催化“新”反应。 特定目标2重点介绍烯醇酶的mandelate Racemase子组的不同成员 超家族（在第七和第六个p链的末端的酸/碱基his-asp dyad 由操纵子编码的结构域），这些域也编码藻酶，脱氢酶，肌动酶， 和/或激酶。 特定的目标3专注于类似Rubisco的蛋白质（RLP），这些蛋白质由操纵子编码 编码异构酶，醛糖酶，转酮酶和其他醛糖/酮5-磷酸的同源物利用 酶。 相关性（请参阅说明）； 将功能分配给由基因组编码的完整蛋白质集是一个主要问题。 但是，当解决此问题时，它们在分子，细胞和生物功能中的作用将是 可以确定已知和新颖的小分子干预目标，从而提供了新的目标 治疗设计的方法。该计划项目的重点是开发和实施 集成序列结构策略，以预测 在基因组项目中发现的未表征的蛋白质，从而促进了其功能分配。