Elucidation of reaction mechanism ofthe enzymes involved in the syntbssis of photosynthetic pigments

光合色素合成酶反应机制的阐明

• 批准号: 18570105
• 负责人: FUKUYAMA Keiichi
• 金额: $ 2.51万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2006
• 资助国家: 日本
• 起止时间: 2006 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18570105/
• 关键词: bilin reductase; photosynthesis; X-ray crystallography; chlorophyll; ferredoxin; bilin pigment; γ-glutamyltranspentidase; 光合成色素; 反応中間体; ビリベルジン; 構造性色素; ビリン環元酵素; 表面電荷

Photosynthetic organisms utilize phytobilins, linear tetrapyrrole pigments, for photosynthesis and light sensing. Such organisms develop light harvesting systems to accomplish efficient photosynthesis in their living environments. Red algae and cyanobacteria have giant protein-pigment complexes called phycobilisomes as a light-harvesting system, in which phycobilins (one of phytobilins) are utilized for light-harvesting pigments. Phytobilins are biosynthesized from heme by heme oxygenase and ferredoxin dependent bilin reductases (FDBRs). PcyA, a member of FDBR family, is unique in reducing biliverdin Xiα (BV) to phycocyanobilin by two sequential steps, in which electrons are supplied by ferredoxin. We previously determined the crystal structure of PcyA from cyanobacterium Synechocystis sp. PCC 6803 in complex with BV. To shed light on the molecular mechanism of PcyA reaction, we synthesized the pigment, the product of the first step of PcyA reduction, and determined the crystal structure of PcyA-pigment complex. On the basis of the structure site directed mutagenesis and functional analysis are under way.γ-Glutamyltranspeptidase (GGT) catalyzes the cleavage of such γ-glutamyl compounds as glutathione, and transfer of their γ-glutamyl group to water or to other amino acids and peptides. Azaserine and acivicin are classical and irreversible inhibitors of GGT, but their binding sites and the inhibition mechanisms remained to be defined. We have determined the crystal structures of GGT from Eacherichia coil in complex with azaserine and acivicin at 1.65 A resolution. They form a covalent bond with the Oγ atom of Thr391, the catalytic residue of GGT. Notably, in the azaserine complex the carbonyl of azaserine is attacked by Thr391 to form a tetrahedral intermediate. When acivicin is bound to GGT, a migration of the single and double bonds occurs in its dihydroisoxazole ring.

光合生物利用植物胆素（线性四吡咯色素）进行光合作用和光传感。这些生物体开发光捕获系统以在其生活环境中实现有效的光合作用。红藻和蓝藻具有巨大的蛋白质-色素复合物，称为藻胆体，作为光捕获系统，其中藻胆素（植物胆素之一）用于捕获光的色素。植物胆素是通过血红素加氧酶和铁氧还蛋白依赖性胆素还原酶 (FDBR) 从血红素生物合成的。 PcyA 是 FDBR 家族的成员，其独特之处在于通过两个连续步骤将胆绿素 Xiα (BV) 还原为藻蓝蛋白，其中电子由铁氧还蛋白提供。我们之前确定了蓝藻集胞藻 (Synechocystis sp) 中 PcyA 的晶体结构。 PCC 6803 与 BV 复合。为了阐明PcyA反应的分子机制，我们合成了PcyA还原第一步的产物颜料，并确定了PcyA-颜料复合物的晶体结构。基于该结构的定点突变和功能分析正在进行中。γ-谷氨酰转肽酶（GGT）催化谷胱甘肽等γ-谷氨酰化合物的裂解，并将其γ-谷氨酰基转移至水或其他氨基酸和肽。重氮丝氨酸和阿西维辛是经典的不可逆 GGT 抑制剂，但它们的结合位点和抑制机制仍有待确定。我们以 1.65 A 的分辨率确定了来自 Eacherichia 线圈的 GGT 与重氮丝氨酸和阿西维星复合物的晶体结构。它们与 Thr391（GGT 的催化残基）的 Oγ 原子形成共价键。值得注意的是，在重氮丝氨酸复合物中，重氮丝氨酸的羰基被 Thr391 攻击，形成四面体中间体。当阿西维星与 GGT 结合时，其二氢异恶唑环中发生单键和双键的迁移。

项目成果

Structures of phytobilin synthesis enzymes

植物胆素合成酶的结构

• 发表时间: 2007
• 作者: K.;Wada;J.;Hiratake;M.;Irie;T.;Okada;C.;Yamada;H.;Kumagai;H.;Suzuki;K.;Fukuyama;K. Fukuyama
• 通讯作者: K. Fukuyama

Crystal Structures of Escherichia coil γ-Glutamyltranspeptidase in Complex with Azaserine and Acivicin : Novel Mechanistic Implication for Inhibition by Glutamine Antagonists

埃希氏菌 γ-谷氨酰转肽酶与重氮丝氨酸和 Acivicin 复合物的晶体结构：谷氨酰胺拮抗剂抑制的新机制意义

• 期刊: J. Mol. Biol (in press)
• 作者: K.;Wada;J.;Hiratake;M.;Irie;T.;Okada;C.;Yamada;H.;Kumagai;H.;Suzuki;K.;Fukuyama
• 通讯作者: Fukuyama

Mass spectroscopic identification of lysine residues of heme oxygenase-1 that are involved in its interaction with NADPH-cytochrome P450 reductase

质谱鉴定参与其与 NADPH-细胞色素 P450 还原酶相互作用的血红素加氧酶 1 的赖氨酸残基

• 发表时间: 2008
• 期刊: Biochem. Biophys. Res. Commun 367
• 作者: Y.;Higashimoto;M.;Sugishima;H.;Sato;H.;Sakamoto;K.;Fukuyama;G.;Palmer;M.;Noguchi
• 通讯作者: Noguchi

X-Ray Crystallographic and Biochemical Characterization of the lnhibitory Action of an lmidazole-dioxolate Compound on Heme Oxygenase

咪唑二氧醇化合物对血红素加氧酶抑制作用的 X 射线晶体学和生化表征

• 发表时间: 2007
• 期刊: Biochemistry 46
• 作者: A.;Dey;F.E.;Jenney;Jr.;M.W.W.;Adams;E.;Babini;Y.;Takahashi;K.;Fukuyama;K.O.;Hodgson;B.;Hedman;E.I.;Solomon;M. Sugishima
• 通讯作者: M. Sugishima

Crystal structures of BchU, a methyltransferase involved in bacteriochlorophyll c biosynthesis, and its complex with S-adenosylhomocysteine: implications for reaction mechanism.

• DOI: 10.1016/j.jmb.2006.05.057
• 发表时间: 2006-07
• 期刊: Journal of molecular biology
• 影响因子: 5.6
• 作者: K. Wada;H. Yamaguchi;J. Harada;Keiko Niimi;S. Osumi;Y. Saga;H. Oh-oka;H. Tamiaki;K. Fukuyama