MECHANISMS OF PYRIDOXAL 5' PHOSPHATE DEPENDENT ENZYMES

含有吡哆醛-5‘-磷酸(PLP)的酶，也被称为维生素B6， 在生物学中普遍存在，在新陈代谢中执行必要的反应 氨基酸和胺。这些酶催化多种不同的 反应，包括外消旋，转氨化，α-和β- 脱羧化，逆转羟醛裂解，β和伽马消除和 换人。PLP依赖的酶在代谢中的中心作用 氨基酸和胺的合成以及重要生物活性的生物合成 代谢物，使它们成为机制设计的有吸引力的目标- 基于抑制剂，这可能是有用的药物。碳-碳裂解酶 (酪氨酸苯酚裂解酶和色氨酸吲哚裂解酶)是细菌酶 催化L酪氨酸或L-酪氨酸的水解性β消除 色氨酸分别给出苯酚或吲哚和丙酮酸铵。 这些酶是研究结构之间关系的理想模型。 以及PLP酶的作用机制。它们的结构和机制 酶的研究将通过诱变，X射线结晶学和前 稳态动力学。犬尿氨酸酶催化水解性裂解 L-犬尿氨酸给邻氨基苯甲酸，L-丙氨酸。在真核生物中 生物，犬尿氨酸酶的产物，3-羟基邻氨基苯甲酸， 在双加氧酶的作用下发生环状裂解，生成α-氨基-β-氨基丁酸 羧基多锥半醛，可自发环合得到 喹啉酸，一种已知的神经毒素。犬尿氨酸酶抑制剂可以 因此在治疗一些严重的神经疾病方面有价值， 如亨廷顿舞蹈症、肌萎缩侧索硬化症、中风、癫痫和艾滋病 痴呆症。为了帮助设计新的抑制剂，犬尿氨酸酶 将被克隆，测序，并确定三维结构。 去年，NIFS的产物被发现是一种依赖PLP的酶 催化L-半胱氨酸脱硫制得L-丙氨酸和 元素硫。有人提出，这种酶是需要的 铁硫中心的体外合成，这是必需的 电子传递过程，因此在生物学中无处不在。这个 L-半胱氨酸脱硫酶的机理将用稳态和红外光谱进行研究 准稳态动力学方法。我们继续研究的结果 可能会对改进的目标做出重大贡献 未来美国人的医疗保健。

Enzymes containing pyridoxal-5'-phosphate (PLP), also known as vitamin B6, are ubiquitous in biology, performing essential reactions in metabolism of amino acids and amines. These enzymes catalyse a wide variety of reactions, including racemization, transamination, alpha- and beta- decarboxylation, retro-aldol cleavage, beta- and gamma-elimination and substitution. The central role of PLP-dependent enzymes in the metabolism of amino acids and amines, and the biosynthesis of important bioactive metabolites, makes them attractive targets for the design of mechanism- based inhibitors, which may be useful as drugs. The carbon-carbon lyases (tyrosine phenol-lyase and tryptophan indole-lyase) are bacterial enzymes which catalyse the hydrolytic beta-elimination of L-tyrosine or L- tryptophan to give, respectively, phenol or indole, and ammonium pyruvate. These enzymes are ideal models to study the relationship between structure and mechanisms in PLP enzymes. The structures and mechanisms of these enzymes will be studied by mutagenesis, X-ray crystallography, and pre- steady-state kinetics. Kynureninase catalyses the hydrolytic cleavage of L-kynurenine to give anthranilic acid and L-alanine. In eucaryotic organisms, the product of kynureninase, 3-hydroxyanthranilic acid, undergoes ring cleavage by a dioxygenase to give alpha-amino-beta- carboxymuconic semialdehyde, which can cyclize spontaneously to give quinolinic acid, a known neurotoxin. Inhibitors of kynureninase could thus be of value in the treatment of some serious neurological disorders, such as Huntington's chorea, ALS, stroke, epilepsy, and AIDS-related dementia. In order to aid in the design of new inhibitors, kynureninase will be cloned, sequenced, and the three dimensional structure determined. Last year, the product of nifS was found to be a PLP-dependent enzyme which catalyses the desulfurization of L-cysteine to give L-alanine and elemental sulfur. It has been proposed that this enzyme is required for the in vitro synthesis of iron-sulfur centers, which are necessary for electron transfer processes and are thus ubiquitous in biology. The mechanism of L-cysteine desulfurase will be studied by steady-state and pre-steady-state kinetic methods. The results of our continuing research are likely to make a significant contribution to the goal of improving health care for Americans in the future.