HEME A AND CYTOCHROME OXIDASE BIOSYNTHESIS

血红素 A 和细胞色素氧化酶生物合成

• 批准号: 2022785
• 负责人: ALEXANDER A TZAGOLOFF
• 金额: $ 18.92万
• 依托单位: COLUMBIA UNIV NEW YORK MORNINGSIDE
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-01-15 至 1998-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2022785
• 关键词: Saccharomyces cerevisiae; biopsy; complementary DNA; cytochrome oxidase; enzyme biosynthesis; enzyme deficiency; fungal genetics; heme; human genetic material tag; human tissue; methyl group; microorganism metabolism; mitochondria; molecular cloning; mutant; oxidation; phenotype; porphyrin biosynthesis; protein structure function

Cytochrome oxidase promotes the transfer of electrons from reduced cytochrome c to molecular oxygen, the terminal step of the respiratory chains of mitochondria and aerobic microorganisms. In S. cerevisiae, this membrane complex is composed of 10 different subunit polypeptides of which three are encoded in mitochondrial DNA. The two genetically distinct groups of polypeptides are assembled into the holoenzyme by an elaborate process that requires the expression of some 40 nuclear genes. The products of two such genes, COX10 and COX11, have recently been implicated to function in the synthesis of heme A, a prosthetic group unique to cytochrome oxidase. One of the goals of this proposal is to characterize the roles of the COX10 and COX11 proteins in farnesylation of the 8-vinyl and oxidation of the 2-methyl substituents of the porphyrin ring and to screen for other heme A mutants. These strains will be used to used to identify the intermediates and enzymes of heme A biosynthesis, a pathway about which virtually nothing is known at present. A substantial number of non-maternally inherited human myopathies have been ascribed to decreased levels of cytochrome oxidase in muscle mitochondria. Despite extensive studies, none of these fatal diseases have been correlated with mutations in the structural or catalytic subunits of this respiratory complex. It is not unreasonable to think that in some cases the enzyme deficiency may stem from mutations affecting heme A biosynthesis. To assess this possibility, muscle biopsies from patients with cytochrome oxidase defects will be analyzed for the accumulation of heme A intermediates. This approach has been helpful in identifying a novel heme compound in a cox11 mutant of yeast. Secondly, the human cDNA homologs of COX10 and COX11 will be cloned in order to facilitate a molecular analysis of the comparable cDNAs from myopathic tissues. The cytochrome oxidase deficient phenotype of yeast strains assigned to approximated 11 complementation groups is elicited by mutations affecting late events in the enzyme assembly pathway. The third goal of this proposal is to complete the analysis of this set of genes and to clarify the functions supplied by the encoded proteins. These studies are anticipated to provide a blueprint of the genetic information and the molecular mechanisms governing assembly of the functional complex.

细胞色素氧化酶促进电子从还原态转移 细胞色素c转化为分子氧，呼吸的最后一步 线粒体和需氧微生物的链。 在酿酒酵母中， 该膜复合物由 10 个不同的亚基多肽组成 其中三个是在线粒体 DNA 中编码的。 两者在基因上 不同的多肽组通过以下方式组装成全酶： 复杂的过程需要约 40 个核基因的表达。 两个此类基因 COX10 和 COX11 的产物最近已被 与血红素 A（一种辅基）的合成有关 细胞色素氧化酶所特有的。 该提案的目标之一是 表征 COX10 和 COX11 蛋白在法呢基化中的作用 8-乙烯基的氧化和2-甲基取代基的氧化 卟啉环并筛选其他血红素 A 突变体。 这些菌株 将用于鉴定血红素中间体和酶 一种生物合成，一种目前几乎一无所知的途径 展示。 大量非母系遗传的人类肌病 归因于肌肉中细胞色素氧化酶水平降低 线粒体。 尽管进行了大量研究，这些致命疾病都没有 与结构或催化的突变有关 该呼吸复合体的亚单位。 想想也不无道理 在某些情况下，酶缺乏可能源于突变 影响血红素A的生物合成。 为了评估这种可能性，肌肉 将分析细胞色素氧化酶缺陷患者的活组织检查 用于血红素A中间体的积累。 这种方法已被 有助于鉴定酵母 cox11 突变体中的新型血红素化合物。 其次，COX10和COX11的人类cDNA同源物将被克隆到 为了促进对来自以下来源的可比 cDNA 进行分子分析 肌病组织。 酵母菌株的细胞色素氧化酶缺陷表型分配给 大约 11 个互补组是由影响的突变引起的 酶组装途径中的晚期事件。 本次的第三个目标 建议完成这组基因的分析并澄清 由编码的蛋白质提供的功能。 这些研究是 预计将提供遗传信息和 控制功能复合物组装的分子机制。