STRUCTURE-FUNCTION RELATIONS IN PYRIDINE COENZYMES

我们计划发起一项全面的努力，以期在 基本水平，即吡啶核苷酸的化学。然后我们 建议应用这些知识来理解吡啶的作用机理。 辅酶依赖酶。我们的工具将是传统的物理有机工具 化学研究吡啶核苷酸的化学性质， 设计和合成多种吡啶的生物有机化学 核苷酸类似物来测试我们提出的机制和一种酶 用脱氢酶和双酶联用评价其性质的方法 水解酶。我们将研究这种化学物质的作用机理。 新型NAD类似物中烟酰胺糖链的水解 旨在解决有关酸和碱的特定机理问题 糖基水解的催化作用。我们将利用这些信息来调查 烟酰胺糖基的酶促水解机理 Bond，重点是NAD-糖水解酶和ADP-核糖基转移酶， 包括依赖NAD的霍乱和白喉毒素。我们还将 还原形式的无环化合物的化学和酶性质研究 NAD类似物作为一类潜在的底物依赖的活性物质 脱氢酶的定点失活剂。最后，我们将探讨 辅酶-脱氢酶相互作用的立体化学限制 烟酰胺部分，以确定相互作用 负责立体定向氢化物转移的保真度。这个 调查将使我们能够更深入地了解 这类极其重要的辅酶的功能和机制 酶对它们起作用的方式。

We plan to launch a comprehensive effort toward understanding, at a fundamental level, the chemistry of the pyridine nucleotides. We then propose to apply this knowledge to understand the mechanisms of pyridine coenzyme-dependent enzymes. Our tools will be traditional physical organic chemistry to study the chemical properties of pyridine nucleotides, bioorganic chemistry to design and synthesize a wide range of pyridine nucleotide analogues to test our proposed mechanisms and an enzymological approach to evaluate their properties with both dehydrogenases and hydrolytic enzymes. We will investigate the mechanisms of the chemical hydrolysis of the nicotinamide glycosyllinkage in new NAD+ analogues designed to address specific mechanistic questions regarding acid and base catalysis of glycosyl hydrolysis. We will use this information to probe the mechanism of the enzymatic hydrolysis of the nicotinamide glycosyl bond, focusing on the NAD-glycohydrolases and ADP-ribosyltransferases, including the NAD+-dependent cholera and diphtheria toxin. We will also study the chemical and enzymatic properties of reduced forms of acyclic NAD+ analogues as a potential new class of substrate-dependent, active site-directed inactivators of dehydrogenases. Finally we will explore the stereochemical constraints of the coenzyme-dehydrogenase interaction with the nicotinamide moiety in order to determine the interactions that are responsible for the fidelity of stereospecific hydride transfer. The investigations will allow us to achieve a deeper understanding of the functioning of this vitally important class of coenzymes and the mechanism by which enzymes act upon them.