Studies on Enzyme Mechanism and Inhibitor Design

本提案分为三节。第一部分描述了我们对产生生物碱的酶的研究。生物碱是主要由植物和真菌产生的含氮天然产物。它们通常具有有效的生物活性，许多已被用作有价值的药物（吗啡/疼痛，长春碱/癌症，麦角胺/偏头痛）。我们正在研究与麦角胺等吲哚生物碱生物合成有关的酶。我们发现这些酶可以产生新的生物碱，可能具有独特的生物学特性。通过了解这些酶如何发挥作用的细节，我们可能能够利用它们来生产新的生物活性化合物。第二部分涉及对导致溃疡和食物中毒的病原菌中甲基萘醌（维生素K2）的生物合成的研究。这些细菌需要甲萘醌进行呼吸，没有它它们就无法生存。我们正在调查的酶参与甲基萘醌在这些生物体的生物合成，因为这些是有吸引力的抑制剂设计的目标。这种抑制剂会阻止酶的运作，因此可以作为抗生素来对抗这些生物体。在我们最初的研究中，我们概述了形成这种维生素的正确生物合成途径，在正在进行的工作中，我们正在研究该途径中的单个酶，以便设计抑制剂。第三部分是关于微管蛋白修饰酶的研究。微管形成人体细胞的细胞骨架，是细胞正常分裂所必需的。因此，干扰微管正常形成的化合物可作为有效的抗癌剂（长春碱、紫杉醇）。微管是由微管蛋白的自发自组装产生的。我们正在研究一组化学修饰微管蛋白的酶，从而控制其物理性质。我们正在制备能够干扰这些酶作用的抑制剂分子，我们预计这些化合物将具有细胞毒性。这可能成为治疗癌症的新方法。

This proposal is divided into three sections. The first section describes our studies on enzymes that produce alkaloids. Alkaloids are nitrogen-containing natural products primarily produced by plants and fungi. They often possess potent biological activities and many have been used as valuable pharmaceuticals (morphine/pain, vinblastine/cancer, ergotamine/migraines). We are studying the enzymes involved in the biosynthesis of the indole alkaloids such as ergotamine. We have discovered that these enzymes can produce novel alkaloids that may have unique biological properties. By understanding the details of how these enzymes function, we may be able to utilize them in the production of new bioactive compounds. The second section involves studies on the biosynthesis of menaquinone (vitamin K2) in the pathogenic bacteria that are responsible for ulcers and food poisoning. These bacteria require menaquinone for respiration and without it they cannot survive. We are investigating the enzymes involved in menaquinone biosynthesis in these organisms since these are attractive targets for inhibitor design. Such inhibitors would prevent the enzymes from operating and would therefore serve as antibiotics against these organisms. In our initial studies we outlined the correct biosynthetic pathway for the formation of this vitamin, and in ongoing work we are studying individual enzymes in the pathway so that inhibitors can be designed. The third section involves studies on tubulin-modifying enzymes. Microtubules form the cytoskeleton of human cells and are absolutely required for proper cell division. For this reason, compounds that interfere with proper microtubule formation act as potent anticancer agents (vinblastine, taxol). Microtubules are generated by the spontaneous self-assembly of tubulin. We are studying a group of enzymes that chemically modify tubulin and therefore control its physical properties. We are preparing inhibitor molecules that will interfere with the action of these enzymes and we anticipate that such compounds will be cytotoxic. This could potentially serve as a new approach towards treating cancer.