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榛树(欧洲榛Corylus avellana L. Gasaway)3-羟基-3-甲基戊二酰辅酶A还原酶编码基因的分子克隆与功能分析

Molecular cloning and functional analysis of the gene encoding 3-hydroxy-3-methylglutaryl coenzyme A reductase from hazel (Corylus avellana L. Gasaway).

作者信息

Wang Yechun, Guo Binhui, Zhang Fei, Yao Hongyan, Miao Zhiqi, Tang Kexuan

机构信息

Plant Biotechnology Research Center, School of Agriculture and Biology, School of Life Science and Technology, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200030, PR China.

出版信息

J Biochem Mol Biol. 2007 Nov 30;40(6):861-9. doi: 10.5483/bmbrep.2007.40.6.861.

Abstract

The enzyme 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR; EC1.1.1.34) catalyzes the first committed step of isoprenoids biosynthesis in MVA pathway. Here we report for the first time the cloning and characterization of a full-length cDNA encoding HMGR (designated as CgHMGR, GenBank accession number EF206343) from hazel (Corylus avellana L. Gasaway), a taxol-producing plant species. The full-length cDNA of CgHMGR was 2064 bp containing a 1704-bp ORF encoding 567 amino acids. Bioinformatic analyses revealed that the deduced CgHMGR had extensive homology with other plant HMGRs and contained two transmembrane domains and a catalytic domain. The predicted 3-D model of CgHMGR had a typical spatial structure of HMGRs. Southern blot analysis indicated that CgHMGR belonged to a small gene family. Expression analysis revealed that CgHMGR expressed high in roots, and low in leaves and stems, and the expression of CgHMGR could be up-regulated by methyl jasmonate (MeJA). The functional color assay in Escherichia coli showed that CgHMGR could accelerate the biosynthesis of beta-carotene, indicating that CgHMGR encoded a functional protein. The cloning, characterization and functional analysis of CgHMGR gene will enable us to further understand the role of CgHMGR involved in taxol biosynthetic pathway in C. avellana at molecular level.

摘要

3-羟基-3-甲基戊二酰辅酶A还原酶(HMGR;EC1.1.1.34)催化甲羟戊酸(MVA)途径中类异戊二烯生物合成的第一步。在此,我们首次报道了从产紫杉醇的植物榛(Corylus avellana L. Gasaway)中克隆并鉴定了一个编码HMGR的全长cDNA(命名为CgHMGR,GenBank登录号EF206343)。CgHMGR的全长cDNA为2064 bp,包含一个1704 bp的开放阅读框,编码567个氨基酸。生物信息学分析表明,推导的CgHMGR与其他植物HMGR具有广泛的同源性,包含两个跨膜结构域和一个催化结构域。预测的CgHMGR三维模型具有HMGRs典型的空间结构。Southern杂交分析表明CgHMGR属于一个小基因家族。表达分析表明,CgHMGR在根中高表达,在叶和茎中低表达,并且茉莉酸甲酯(MeJA)可以上调CgHMGR的表达。在大肠杆菌中的功能颜色测定表明,CgHMGR可以加速β-胡萝卜素的生物合成,表明CgHMGR编码一种功能蛋白。CgHMGR基因的克隆、鉴定和功能分析将使我们能够在分子水平上进一步了解CgHMGR在榛树紫杉醇生物合成途径中的作用。

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