MOLYBDENUM COFACTOR--BIOSYNTHESIS AND FUNCTION

钼辅因子--生物合成和功能

• 批准号: 6498140
• 负责人: HERMANN SCHINDELIN
• 金额: $ 17.95万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-03-01 至 2003-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6498140
• 关键词: X ray crystallography; biosynthesis; biotin; chemical structure function; cofactor; dimethylsulfoxide; enzyme activity; lyase; molybdenum; nutrition related tag; oxidoreductase; oxidoreductase inhibitor; pteridines; site directed mutagenesis; sulfoxide; tissue /cell culture

The molybdenum cofactor (Moco) is present in four different enzyme families which catalyze important transformations in the global carbon, sulfur and nitrogen cycles. Moco consists of a mononuclear molybdenum coordinated to the sulfur atoms of a dithiolene group which is part of a substituted pterin derivative termed molybdopterin. Enzymes containing this cofactor are found in all three phylogenetic kingdoms of life, and biosynthesis of the cofactor follows the same pathway, both in prokaryotes and in eukaryotes including plants and human. Moco deficiency is inherited as an autosomal recessive trait and is a severe disease in humans which manifests itself in neurological abnormalities leading to premature death in the affected individuals. The experiments outlined in this proposal address two important aspects of Moco, namely its biosynthesis and its detailed function once it has been incorporated into the target enzymes. The specific aims presented in this proposal therefore are: (i) Characterization of the early steps in Moco biosynthesis through structural studies on the enzyme MoaC. MoaC most likely either catalyzes the first step in Moco biosynthesis starting with a phosphorylated guanosine, or the second step in which A molybdopterin precursor is generated which still lacks the sulfur atoms. (ii) Crystal structure analysis of molybdopterin synthase which incorporates the sulfur atoms into a molybdopterin precursor generating molybdopterin. The small subunit of the heterodimeric molybdopterin synthase contains an unusual C-terminal thiocarboxylate which serves as the sulfur donor during catalysis. (iii) Crystal structure determination of MogA which might function as a molybdochelatase and thus would be responsible for incorporating molybdenum, which is present in the cell as molybdate, into the molybdopterin. (iv) High resolution structure analysis of the Moco-containing enzyme dimethylsulfoxide reductase including characterization of different oxidation states and complexes of the enzyme with substrates and inhibitors which will lead to a detailed characterization of the catalytic mechanism. (v) Crystal structure analysis of biotin sulfoxide reductase, which is related in sequence to dimethylsulfoxide reductase but differs in substrate specificity. Comparative studies of the two enzymes will lead to a more general understanding of the roles Moco is playing in enzymes containing this cofactor.

钼辅因子 (Moco) 存在于四种不同的酶中 催化全球碳的重要转变的家庭， 硫和氮循环。 Moco 由单核钼组成 与二硫杂环戊烯基团的硫原子配位，该基团是 称为钼蝶呤的取代蝶呤衍生物。 酶 在所有三个系统发育界中都发现含有该辅因子 生命的过程和辅因子的生物合成遵循相同的途径，两者 在原核生物和真核生物中，包括植物和人类。 莫科 缺陷是作为常染色体隐性遗传的，是一种严重的 人类疾病，表现为神经系统异常 导致受影响个体过早死亡。 实验 该提案概述了 Moco 的两个重要方面，即 它的生物合成及其掺入后的详细功能 转化为目标酶。本提案提出的具体目标 因此： (i) Moco 早期步骤的特征 通过酶 MoaC 的结构研究进行生物合成。 莫阿克最 可能催化 Moco 生物合成的第一步 与磷酸化鸟苷，或第二步，其中A 生成的钼蝶呤前体仍缺乏硫原子。 (ii) 钼蝶呤合酶的晶体结构分析 将硫原子合并到钼蝶呤前体中，产生 钼蝶呤。 异二聚体钼蝶呤的小亚基 合酶含有一种不寻常的 C 末端硫代羧酸盐，可用作 催化过程中的硫供体。 (三)晶体结构 测定可能充当钼螯合酶的 MogA 以及 因此将负责掺入钼，这是存在的 在细胞中以钼酸盐的形式转化为钼蝶呤。 (四) 高分辨率 含Moco酶二甲亚砜的结构分析 还原酶，包括不同氧化态的表征和 酶与底物和抑制剂的复合物将导致 催化机制的详细表征。 (五) 水晶 生物素亚砜还原酶的结构分析，与 与二甲亚砜还原酶序列相同，但底物不同 特异性。 对这两种酶的比较研究将带来更多的结果 对 Moco 在含有酶的酶中发挥的作用的一般了解 这个辅助因子。