Coenzyme A Analogs as Probes of Enzymatic Reactions

这个项目将使用合成辅酶A （CoA）类似物来研究酶催化CoA酯反应的基本机理问题。这项工作是基于在P.I.开发的方法为了方便地合成CoA/CoA酯类似物。最大的努力将是探索烯酸酯和/或烯醇中间体在乙酰辅酶a和其他辅酶a酯的酶催化反应中的作用。制备稳定的烯酸酯和烯醇型乙酰辅酶a的类似物，并研究其作为柠檬酸合酶抑制剂的作用。这些同位素富集形式的类似物与酶结合的质子化状态将通过核磁共振和某些情况下的红外光谱进行研究。类似的研究将与相关的酶进行，包括苹果酸合成酶、巴豆酮酶和乙酰辅酶a依赖的乙酰转移酶。该项目有望为进一步了解辅酶a酯的酶催化反应和酶反应机理的一般原理提供新的见解。德鲁哈默博士将研究需要辅酶A才能起作用的酶的反应。酶是生物系统中进行化学反应的催化剂。了解这些酶的功能是在基本化学水平上理解生物过程的核心。大约4%的酶需要辅酶A （CoA）来维持其催化活性。辅酶a在这些酶促反应中充当反应物或反应物载体。在生物系统中，辅酶a也是通过一系列酶催化反应合成的。在本实验室之前的工作中，开发了一种方法，利用制造辅酶a的酶来制造天然辅酶a的改变形式（类似物）。本研究将利用该方法制备一系列辅酶a类似物，作为研究辅酶a利用酶反应的探针。这将有助于揭示酶如何形成化学键，从简单的构建块制造复杂的生物分子，以及酶如何分解食物中的复杂化合物以产生生物能。＊＊＊

9321371 Drueckhammer This project will use synthetic Coenzyme A (CoA) analogs to study fundamental mechanistic questions in enzyme catalyzed reactions of CoA esters. This work is based on methodology developed in the P.I.'s lab for the convenient synthesis of CoA/CoA ester analogs. The largest effort will be directed at probing the involvement of enolate and/or enol intermediates in enzyme catalyzed reactions of acetyl-CoA and other CoA esters. Stable analogs of the enolate and enol forms of acetyl-CoA will be prepared, and studied as inhibitors of citrate synthase. The protonation state of these analogs in isotopically enriched form bound to the enzyme will be studied by NMR and in some cases FTIR. Similar studies will be conducted with related enzymes, including malate synthase, crotonase, and acetyl-CoA dependent acetyltransferases. This project is expected to provide new insights into the enzyme catalyzed reactions of CoA esters and general principles of enzyme reaction mechanisms. %%% Dr. Drueckhammer will study the reactions of enzymes which require Coenzyme A for their function. Enzymes are catalysts which carry out chemical reactions in biological systems. Understanding how these enzymes function is at the heart of understanding biological processes at a fundamental chemical level. About 4% of all enzymes require coenzyme A (CoA) for their catalytic activity. CoA serves as a reactant or a reactant carrier in these enzymatic reactions. CoA is also synthesized in biological systems by a series of enzyme catalyzed reactions. In previous work in this lab, a method was developed for using the enzymes which make CoA to make altered forms (analogs) of natural CoA. This research will use this method to make a series of CoA analogs designed to serve as probes for studying the reactions of CoA utilizing enzymes. This will help reveal how enzymes form chemical bonds to make complex biological molecules from simple building blocks, and how enzymes break down com plex compounds in food to produce biological energy. ***