FIRST STEP OF HEME BIOSYNTHESIS IN MAMMALS

长期目标是了解5- 氨基乙酰丙酸合酶（ALAS），在酶和基因 水平，在调节血红素生物合成的分化 红细胞 ALAS催化第一步和限速步骤， 血红素的生物合成，甘氨酸的PLP依赖反应， 琥珀酰辅酶A，得到氨基乙酰丙酸、辅酶A和CO2。 唉 合成虽然普遍存在，但主要发生在 红细胞和肝细胞，其中需求更大，因为 血红蛋白和细胞色素P-450的合成。 ALAS的红系特异性同种型（ALAS-E）表达 伴随着细胞的分化和成熟， 红系细胞 最近，一组杂合子点突变 在ALAS-E酶的催化结构域中， 导致人类遗传性疾病X-连锁铁粒幼细胞性贫血 （XLSA）。 ALAS的分子机制的表征- 催化的反应和调控的表达， ALAS-E基因对于设计XLSA的改进疗法至关重要 患者和/或改善对高危家庭成员的诊断。 预计未来四年的研究将利用化学， 生物化学、物理和分子生物学方法， 解决三个主要的具体目标：i）确定K313的作用， 哺乳动物类红细胞ALAS催化反应; ii）鉴定 PLP中甘氨酸环的功能重要残基 ALAS的辅因子结合位点;和iii）评估转录 哺乳动物红系ALAS（ALAS）基因的调控。 这些研究的结果将提供第一个表征 在分子水平上研究ALAS的功能和机制， 将阐明表达的调控机制， 分化红细胞中的ALAS-E。 重要的是，他们会 为解释已经存在的 确定了与XLSA相关的ALAS-E基因突变。

The long term objective is to understand the role of 5- aminolevulinate synthase (ALAS), both at the enzyme and gene levels, in the regulation of heme biosynthesis in differentiating erythrocytes. ALAS catalyzes the first and rate limiting step in heme biosynthesis, the PLP-dependent reaction of glycine and succinyl-CoA to yield aminolevulinic acid, CoA and CO2. ALAS synthesis, although ubiquitous, occurs predominately in erythrocytes and hepatocytes, where demands are greater because of the synthesis of hemoglobin and cytochrome P-450, respectively. The erythroid-specific isoform of ALAS (ALAS-E) is expressed concomitantly with the differentiation and maturation of the erythroid cells. Recently a heterozygous group of point mutations in the catalytic domain of the ALAS-E enzyme has been found to cause the human genetic disorder X-linked sideroblastic anemia (XLSA). Characterization of the molecular mechanisms of the ALAS- catalyzed reaction and of the regulation of the expression of the ALAS-E gene are essential to design improved therapies for XLSA patients and/or for improved diagnosis for at-risk family members. Studies projected for the next four years will utilize chemical, biochemical, physical and molecular biological approaches to address three major specific aims: i) to define the role of K313 in the mammalian erythroid ALAS-catalyzed reaction; ii) to identify the functionally important residues of the glycine loop in the PLP cofactor binding site of ALAS; and iii) to evaluate transcriptional regulation of the mammalian erythroid ALAS (ALAS) gene. Results from these studies will provide the first characterization of the function and mechanism of ALAS at the molecular level and will elucidate the regulatory mechanism(s) of expression of the ALAS-E in differentiation erythrocytes. Significantly, they will provide the framework for the interpretation of the already identified ALAS-E genetic mutations associated with XLSA.