5-Aminolevulinate Synthase and Heme Biosynthesis

描述（由申请人提供）： 5-氨基乙酰丙酸合酶（ALAS）是α-氧代胺合酶家族的吡哆醛5 '-磷酸（PLP）依赖性酶，催化哺乳动物血红素生物合成途径的第一步和调节步骤。编码红系ALAS同种型的基因突变引起X连锁铁粒幼细胞性贫血（XLSA），这是一种导致组织铁水平升高的红细胞生成障碍。开发对XLSA具有普遍疗效的基于吡哆醇的疗法的前景依赖于对ALAS机制、辅因子结合口袋（PLP-fold）及其关系的了解。这是活性位点的设计，需要PLP辅因子-蛋白质相互作用，区分广泛的PLP依赖性酶催化反应中的一种类型的反应。P.I.的最新研究1996年至2002年，美国科学院的实验室对ALAS的活性部位和机制的结构进行了研究，为以下3个假设的检验奠定了基础： 1. PLP辅因子化学的调节控制ALAS的酶机制。 2. ALAS的底物特异性可以通过对PLP依赖性酶的ctoxoaminesynthase家族的蛋白质支架进行微小修饰来获得，其具有相同的一般PLP结合折叠和催化化学。 3. ALAS和谷氨酸l-半醛氨基变位酶（GSA-AT）的不同催化化学，这两种PLP依赖性酶都产生5-氨基乙酰丙酸（ALA）并在两种天然ALA生物合成途径（例如，在动物和植物中），可以通过初级蛋白质支架的“进化”产生。 拟议的研究提供了一种新的策略，建立ALAS和其他α-氧代胺合酶的结构/功能关系。此外，拟议的研究提供了一种新的方法来理解如何改组蛋白质结构域改变功能和酶活性位点如何演变;这种策略有望成为一个关键，特别是确定人类基因组中未识别基因的功能。

DESCRIPTION (provided by applicant): 5-Aminolevulinate synthase (ALAS), a pyridoxal 5'-phosphate (PLP)-dependent enzyme of the alpha-oxoaminesynthase family, catalyzes the first and regulatory step of the mammalian heme biosynthetic pathway. Mutations in the gene encoding the erythroid ALAS isoform cause X-linked sideroblastic anemia (XLSA), an erythropoietic disorder resulting in increased tissue iron levels. The prospect of developing pyridoxine-based therapies of universal efficacy for XLSA relies on the knowledge of the mechanism of ALAS, the cofactor-binding pocket (PLP-fold) and their relationship. It is the design of the active site, entailing the PLP cofactor-protein interaction, that discriminates one type of reaction among the wide gamut of PLP-dependent enzyme-catalyzed reactions. Recent studies in the P.I.'s laboratory on the architecture of the active site and mechanism of ALAS (1996-2002)set the stage for the following 3 hypotheses to be tested: 1. Modulation of the PLP cofactor chemistry controls the enzymatic mechanism of ALAS. 2. Substrate specificity of ALAS can be acquired through minor modifications of the protein scaffold for the ctoxoaminesynthase family of PLP-dependent enzymes, which possesses the same general PLP-binding fold and chemistry of catalysis. 3. The distinct catalytic chemistries of ALAS and glutamate l-semialdehyde aminomutase (GSA-AT), two PLP-dependent enzymes which both produce 5-aminolevulinate (ALA) and play crucial roles in the two natural ALA biosynthetic pathways (e.g., in animals and plants), can be generated by "evolution" of a primary protein scaffold. The proposed studies provide a new strategy to establish structure/function relationships within ALAS and other alpha-oxoamine synthase enzymes. Further, the proposed studies provide a novel approach to understanding how shuffling protein structural domains changes function and how enzyme active sites evolve; this strategy promises to be a key particularly for determining the function of unidentified genes in the human genome.