紫穗槐-4,11-二烯合酶及其代谢工程研究进展  

Recent advances in the study of amorpha-4,11-diene synthase and its metabolic engineering

作　　者：孔建强[1] 黄勇[1] 沈君豪[1] 王伟[1] 程克棣[1] 朱平[1] 

机构地区：[1]中国医学科学院,北京协和医学院药物研究所,卫生部天然药物生物合成重点实验室,中草药物质基础与资源利用教育部重点实验室,北京100050

出　　处：《药学学报》2009年第12期1320-1327,共8页Acta Pharmaceutica Sinica

基　　金：科技部"863"重点项目资助项目(2007AA021501);国家自然科学基金资助项目(30701061);北京市自然科学基金资助项目(7082063);教育部博士点新教师基金资助项目(20070023077);药物所基本科研业务费(2006QN05)资助项目

摘　　要：紫穗槐-4,11-二烯合酶催化FPP(farnesyl pyrophosphate,法尼基焦磷酸)生成青蒿素前体紫穗槐-4,11-二烯,是青蒿素生物合成途径中的关键酶。本文对紫穗槐-4,11-二烯合酶的分子生物学和代谢工程研究进行了综述。紫穗槐-4,11-二烯合酶编码基因及其相关核酸序列已经得到了克隆。紫穗槐-4,11-二烯合酶cDNA全长1641bp,编码546aa。紫穗槐-4,11-二烯合酶最适pH范围较宽,但需要二价金属离子作为辅酶才能发挥作用,其产物和底物的特异性不高。在紫穗槐-4,11-二烯合酶作用下,FPP首先进行的是1,6-合环,然后是1,10-合环,形成紫穗槐-4,11-二烯。由于紫穗槐-4,11-二烯合酶在青蒿素生物合成中具有重要的意义,自从其基因被克隆测序后,先后被导入E.coli、S.cereviseae、烟草、拟南芥和A.nidulans,获得了能产生紫穗槐-4,11-二烯的各种工程菌或细胞,研究通过不同方式提高工程菌中紫穗槐-4,11-二烯产量的方法。Amorpha-4,11-diene synthase （ADS） can convert farnesyl pyrophosphate （FPP） to amorpha-4, 11-diene, a precursor of artemisinin. ADS plays an important role in the biosynthesis of artemisinin. This review summarizes the molecular biology and metabolic engineering study of ADS in recent years. The genomic DNA and its cDNA sequences of amorpha-4, 11-diene synthase were cloned from Artemisia annua L. The cDNA encoding amorpha-4, 11-diene synthase contains a 1 641 bp open reading frame coding for 546 amino acids. ADS shows a broad pH optimum and an absolute requirement for divalent metal ions as cofactors. The specificity of ADS to the substrates and products is not high and the formation of amorpha-4,11-diene by ADS from FPP is achieved by an initial 1, 6-closure with subsequent 1, 10-closure. The ADS cDNA cloned from Artemisia annua L, or totally synthesized by PCR, was introduced into different hosts including E. coli, S. cerevisiae, Nicotiana tabacum L. Arabidopsis thaliana and A. nidulans resulting in varied engineering microorganisms and cells producing amorpha-4, 11-diene. The way to improve the production of amorpha4, 11-diene was investigated by two strategies such as improving the supply of substrate and directing FPP flux to amorpha-4, 11-diene production from competing pathways.

关 键 词：紫穗槐-4 11-二烯 11-二烯合酶 青蒿素 工程菌 

分 类 号：R915.1[医药卫生—微生物与生化药学]