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Molecular Mechanism of the Catalytic Function of prenyl Chain Elongating Enzymes Participating in the biosynthesis of Isoprenoid Compounds

异戊二烯基链延长酶参与类异戊二烯化合物生物合成催化作用的分子机制

基本信息

批准号：
12480169
负责人：
KOYAMA Tanetoshi
金额：
$ 8.77万
依托单位：
TOHOKU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2002
项目状态：
已结题

项目摘要

In the biosynthesis of isoprenoid compounds all of the carbon backbones are derived from prenyl diphosphates, which are synthesized by the action of prenyltransferases.During the last decades the structural genes for many kinds of prenyltransferases catalyzing (E)-type prenyl chain elongation have been identified and characterized. However, no information has been available about the structures of (Z)-prenyl diphosphate syntheses until our isolation of the gene for Micrococcus luteus undecaprenyl diphosphate synthase in 1998. The amino acid sequence of the (Z)-prenyl diphosphate synthase is totally different from those of (E)-prenyl chain elongating enzymes.The crystal structure of the undecaprenyl diphosphate synthase has been determined at 2.2 A resolution as the first three dimensional structure among cis-prenyl chain elongating enzymes. This enzyme shows a novel protein fold which is completely different from those for the enzymes relating to isoprenoid biosynthesis, having a commo … More n structure "isoprenoid synthase fold". A plausible substrate-binding model of the enzyme has been deduced from the precise three-dimensional structure as well as the data accumulated from site-directed mutagenesis experiments.We have isolated and characterized the cDNA for the rubber transferaes from Hevea brasiliensis. In viro rubber production using the recombinant enzyme overexpressed in E. coli cells revealed that the Hevea rubber transferase might require some activation factors in the washed bottom particle fraction of Hevea latex for the production of high molecular weight rubber molecules.Dolichol has an essential role in the biosynthesis of N-linked glycoproteins and glycosylphosphatidylinositol-anchored proteins as the glycosyl carrier lipid in higher organisms. Through the database search for a gene encoding highly conserved regions for cis-prenyl chain elongating enzymes, we have identified a human dehydrodolichyl diphosphate synthase. The mRNA expression levels of the dehydrodolichyl diphosphate synthase in human tissues were analyzed by Northern blotting to show high expressions in testis as well as in kidney. Less

在类异戊二烯化合物的生物合成中，所有的碳骨架都来自于异戊二烯基二磷酸，而异戊二烯基二磷酸是在异戊二烯基转移酶的作用下合成的，在过去的几十年中，许多催化（E）型异戊二烯基链延长的异戊二烯基转移酶的结构基因已经被鉴定和表征。然而，直到我们在1998年分离出藤黄微球菌十一异戊二烯基二磷酸合酶的基因，才有关于（Z）-异戊二烯基二磷酸合成酶结构的信息。（Z）-异戊烯基二磷酸合酶的氨基酸序列与（E）-异戊烯基链延长酶完全不同，十一异戊烯基二磷酸合酶的晶体结构在2.2 A分辨率下被确定，是顺式异戊烯基链延长酶中第一个三维结构。这种酶显示了一种新的蛋白质折叠，它与类异戊二烯生物合成相关酶的蛋白质折叠完全不同， ...更多信息 n结构“类异戊二烯合成酶折叠”。从精确的三维结构以及定点突变实验积累的数据中推导出了一个合理的底物结合模型。我们分离并鉴定了橡胶树橡胶转移酶的cDNA。利用在E.大肠杆菌细胞的研究表明，橡胶树橡胶转移酶可能需要橡胶树胶乳洗涤后的底部颗粒组分中的某些活化因子来产生高分子量的橡胶分子，而Dolichol作为糖基载体脂质在高等生物的N-连接糖蛋白和糖基磷脂酰肌醇锚定蛋白的生物合成中起着重要作用。通过数据库搜索编码顺式异戊二烯基链延长酶的高度保守区域的基因，我们已经确定了人类脱氢二磷酸合酶。通过北方印迹分析人组织中脱氢二磷酸二氢胆甾醇合酶的mRNA表达水平，以显示在睾丸和肾脏中的高表达。少