植物萜类合酶研究进展

随着植物中许多有价值的萜类化合物被发现和应用于人类生活 ,萜类生物合成途径的研究倍受重视。萜类合酶催化单萜、倍半萜和二萜生物合成 ,即分别催化GPP、FPP和GGPP形成单萜、倍半萜和二萜。本文叙述了近年来在植物萜类合酶催化机理、克隆策略和萜类生物工程的研究进展     

植物萜类合酶研究进展

随着植物中许多有价值的萜类化合物被发现和应用于人类生活,萜类生物合成途径的研究倍受重视。萜类合酶催化单萜、倍半萜和二萜生物合成,即分别催化GPP、FPP和GGPP形成单萜、倍半萜和二萜。本文叙述了近年来在植物萜类合酶催化机理、克隆策略和萜类生物工程的研究进展。     

倍半萜合酶催化机制的研究进展

萜类化合物是自然界中普遍存在的一大类天然产物,其结构多样。倍半萜化合物是萜类化合物的重要组成部分。倍半萜合酶是倍半萜化合物合成过程中的关键酶。本文综述了近年来多种倍半萜合酶的突变研究,阐明了倍半萜合酶的催化机理。     

青蒿倍半萜合酶(环化酶)研究进展

青蒿素是从中药青蒿中分离得到的抗疟有效单体,是含有过氧基团的新型倍半萜内酯化合物,是目前世界上最有效的疟疾治疗药物。青蒿素的生物合成途径属于类异戊二烯代谢途径中的倍半萜类分支途径,倍半萜合酶是该途径的关键酶之一,目前已从青蒿中克隆了多个倍半萜合酶基因。综述了青蒿中已克隆的几种倍半萜合酶基因的研究进展。     

青蒿倍半萜合酶(环化酶)研究进展

青蒿素是从中药青蒿中分离得到的抗疟有效单体,是含有过氧基团的新型倍半萜内酯化合物,是目前世界上最有效的疟疾治疗药物。青蒿素的生物合成途径属于类异戊二烯代谢途径中的倍半萜类分支途径,倍半萜合酶是该途径的关键酶之一,目前已从青蒿中克隆了多个倍半萜合酶基因。综述了青蒿中已克隆的几种倍半萜合酶基因的研究进展。     

植物萜类生物合成中的后修饰酶

萜类化合物由于其结构类型丰富多样而被称为"terpenome".除了参与植物生长发育、环境应答等生理过程,萜类化合物还应用于医药、有机化工等领域.萜类的生物合成大致可分为前体形成、骨架构建以及后修饰三部分,基本骨架通常由萜类合酶催化形成,进一步在后修饰酶的作用下产生数以万计的萜类化合物.结合我们对香茶菜二萜生物合成的初步研究结果,本文主要针对近年来植物萜类生物合成中的一些有代表性的后修饰酶包括P450单氧酶、双键还原酶、酰基转移酶和糖基转移酶,进行研究现状分析与展望.     

谷氨酸棒状杆菌异源合成萜类化合物的研究进展

萜类化合物具有可观的商业价值,但生产过程复杂,产量低,利用微生物异源合成萜类化合物已成为热点。谷氨酸棒状杆菌内含合成萜类色素的途径,具有异源合成萜类化合物的天然优势和研究前景。首次对谷氨酸棒状杆菌合成萜类化合物进行了综述,从萜类合成途径、关键酶和全局调控机制三个方面进行了途经介绍。概述了谷氨酸棒状杆菌中单萜、倍半萜、四萜类化合物的异源合成,并对利用谷氨酸棒状杆菌高效合成萜类化合物所需解决的问题进行讨论,为谷氨酸棒状杆菌高效合成萜类化合物提供建议。     

植物类萜生物合成途径及关键酶的研究进展

萜类化合物是植物中广泛存在的一类代谢产物,在植物的生长、发育过程中起着重要的作用。植物中的萜类化合物有两条合成途径:甲羟戊酸途径和5-磷酸脱氧木酮糖/2C-甲基4-磷酸-4D-赤藓糖醇途径。这两条途径中都存在一系列调控萜类化合物生成、结构和功能各异的酶,其中关键酶的作用决定了下游萜类化合物的产量。植物类萜生物合成途径的调控以及该途径中关键酶的研究已成为目前国内外生物学领域的一大热点。综述了植物类萜生物合成途径和参与该途径的关键酶及其基因工程的研究进展,并展望了其应用前景。     

温郁金萜类合酶基因CwTPS4的克隆和表达特征

温郁金是著名的"浙八味"之一,药用价值高、应用广泛,萜类化合物是其主要的药用成分.萜类合酶是植物萜类化合物生物合成途径中的关键酶.依据温郁金根茎的转录组数据,采用反转录PCR获得了1个萜类合酶基因CwTPS4(GenBank登录号:MW774935),其开放阅读框长1515 bp,编码504个氨基酸,含有萜类合酶特有的结构域和保守序列RXR、DDXXD等;生物信息学分析表明CwTPS4编码的蛋白定位在细胞质中、无跨膜区域,为水溶性的稳定蛋白,属于萜类合酶的TPS-a亚家族成员.实时荧光定量分析表明,CwTPS4基因主要在生长旺盛的叶片中表达,其次在根茎膨大初期的根茎中表达量较高,而在成熟或衰老的组织中表达量较低.本研究从温郁金中克隆得到1个新的萜类合酶基因CwTPS4,经生物信息学分析表明CwTPS4可能参与了温郁金中倍半萜类化合物的合成,为下一步研究其在温郁金萜类物质合成中的功能奠定了一定的基础.     

植物三萜皂苷生物合成中关键后修饰酶研究进展

三萜皂苷是由三萜苷元、糖基、糖醛酸等组成的C30萜类化合物,是许多药用植物的主要活性成分,具有广泛的药理作用。三萜皂苷的生物合成包括前体和三萜皂苷骨架的形成以及调控皂苷结构多样性的后修饰。三萜皂苷的后修饰包括三萜骨架的氧化/羟基化和糖基化,分别由不同超基因家族编码的细胞色素P450单加氧酶和糖基转移酶进行催化。三萜皂苷通过后修饰最终可形成多种单体皂苷。目前,已在少数植物中识别和确认了个别与三萜皂苷生物合成相关的关键后修饰酶,发现了部分很可能参与后修饰过程的候选基因。该文就近年来国内外有关三萜皂苷生物合成途径关键后修饰酶的研究进行综述,为进一步开展相关研究和对合成精细途径的解析提供参考。     

植物萜类化合物的生物合成及应用

萜类化合物是植物中广泛存在的一类代谢产物,在植物生长、发育过程中起重要作用。植物中的萜类化合物有2条合成途径,即甲羟戊酸途径和甲基赤藓糖醇磷酸途径。这2条途径中都存在一系列调控萜类化合物生成、结构和功能各异的酶。植物萜类化合物不仅在植物生命活动中起重要作用,而且具有重要的商业价值,被广泛用于工业、医药卫生等领域。     

Catabolism of citronellol and related acyclic terpenoids in pseudomonads

Terpenes are a huge group of natural compounds characterised by their predominantly pleasant smell. They are built up by isoprene units in cyclic or acyclic form and can be functionalised by carbonyl, hydroxyl or carboxyl groups and by presence of additional carbon–carbon double bonds (terpenoids). Currently, much more than 10,000 terpenoid compounds are known, and many thereof are present in different iso- and stereoforms. Terpenoids are secondary metabolites and can have important biological functions in living organisms. In many cases, the biological functions of terpenoids are not known at all. Nevertheless, terpenoids are used in large quantities as perfumes and aroma compounds for food additives. Terpenoids can be also precursors and building blocks for synthesis of complex chiral compounds in chemical and pharmaceutical industry. Unfortunately, only few terpenoids are available in large quantities at reasonable costs. Therefore, characterisation of suited biocatalysts specific for terpenoid compounds and development of biotransformation processes of abundant terpenoids to commercially interesting derivates becomes more and more important. This minireview summarises knowledge on catabolic pathways and biotransformations of acyclic monoterpenes that have received only little attention. Terpenoids with 20 or more carbon atoms are not a subject of this study.     

Perspectives and limits of engineering the isoprenoid metabolism in heterologous hosts

Terpenoids belong to the largest class of natural compounds and are produced in all living organisms. The isoprenoid skeleton is based on assembling of C5 building blocks, but the biosynthesis of a great variety of terpenoids ranging from monoterpenoids to polyterpenoids is not fully understood today. Terpenoids play a fundamental role in human nutrition, cosmetics, and medicine. In the past 10 years, many metabolic engineering efforts have been undertaken in plants but also in microorganisms to improve the production of various terpenoids like artemisinin and paclitaxel. Recently, inverse metabolic engineering and combinatorial biosynthesis as main strategies in synthetic biology have been applied to produce high-cost natural products like artemisinin and paclitaxel in heterologous microorganisms. This review describes the recent progresses made in metabolic engineering of the terpenoid pathway with particular focus on fundamental aspects of host selection, vector design, and system biotechnology.     

Bacterial diterpene synthases: new opportunities for mechanistic enzymology and engineered biosynthesis

Diterpenoid biosynthesis has been extensively studied in plants and fungi, yet cloning and engineering diterpenoid pathways in these organisms remain challenging. Bacteria are emerging as prolific producers of diterpenoid natural products, and bacterial diterpene synthases are poised to make significant contributions to our understanding of terpenoid biosynthesis. Here we will first survey diterpenoid natural products of bacterial origin and briefly review their biosynthesis with emphasis on diterpene synthases (DTSs) that channel geranylgeranyl diphosphate to various diterpenoid scaffolds. We will then highlight differences of DTSs of bacterial and higher organism origins and discuss the challenges in discovering novel bacterial DTSs. We will conclude by discussing new opportunities for DTS mechanistic enzymology and applications of bacterial DTS in biocatalysis and metabolic pathway engineering.     

Research progress relating to the role of cytochrome P450 in the biosynthesis of terpenoids in medicinal plants

Terpenoids are an extensive and diverse group of plant secondary metabolites. To date, they have been applied in many fields including industry, medicine and health. The wide variety of terpenoid compounds cannot arise solely from simple cyclisations of a precursor molecule or from a single-step reaction; their structural diversity depends on the modification of many specific chemical groups, rearrangements of their skeletal structures and on the post-modification reactions. Most of the post-modification enzymes that catalyse these reactions are cytochrome P450 monooxygenases. Therefore, the discovery and identification of plant P450 genes plays a vital role in the exploration of terpenoid biosynthesis pathways. This review summarises recent research progress relating to the function of plant cytochrome P450 enzymes, describes P450 genes that have been cloned from full-length cDNA and identifies the function of P450 enzymes in the terpenoid biosynthesis pathways of several medicinal plants.     

萜类合成酶定向进化的新思路

萜类化合物是天然产物中种类最多且主要存在于植物和微生物体内的一类化合物。随着越来越多具有应用价值的萜类化合物被挖掘,其应用前景引起了人们的关注,但由于含量低、提取成本高等缺点,因此制约了萜类化合物的广泛应用。合成生物学的兴起,为异源合成具有应用价值的萜类化合物提供了新思路,使构建定向、高效的微生物细胞工厂成为现实。萜类合成酶常作为萜类化合物异源合成代谢调控的靶酶,但天然的萜类合成酶存在催化效率低、底物专一性差、立体/区域选择性差、稳定性差等问题,严重影响萜类化合物的产量。萜类合成酶的定向进化可以有效地解决上述问题,为实现微生物细胞工厂异源、高效合成萜类化合物奠定基础。本文综述了近年来酶的定向进化技术的最新进展及应用,并提出了萜类合成酶定向进化的策略。     

Improving yeast strains using recyclable integration cassettes,for the production of plant terpenoids

Background  

Terpenoids constitute a large family of natural products, attracting commercial interest for a variety of uses as flavours, fragrances, drugs and alternative fuels. Saccharomyces cerevisiae offers a versatile cell factory, as the precursors of terpenoid biosynthesis are naturally synthesized by the sterol biosynthetic pathway.     

Production and engineering of terpenoids in plant cell culture

Terpenoids are a diverse class of natural products that have many functions in the plant kingdom and in human health and nutrition. Their chemical diversity has led to the discovery of over 40,000 different structures, with several classes serving as important pharmaceutical agents, including the anticancer agents paclitaxel (Taxol) and terpenoid-derived indole alkaloids. Many terpenoid compounds are found in low yield from natural sources, so plant cell cultures have been investigated as an alternate production strategy. Metabolic engineering of whole plants and plant cell cultures is an effective tool to both increase terpenoid yield and alter terpenoid distribution for desired properties such as enhanced flavor, fragrance or color. Recent advances in defining terpenoid metabolic pathways, particularly in secondary metabolism, enhanced knowledge concerning regulation of terpenoid accumulation, and application of emerging plant systems biology approaches, have enabled metabolic engineering of terpenoid production. This paper reviews the current state of knowledge of terpenoid metabolism, with a special focus on production of important pharmaceutically active secondary metabolic terpenoids in plant cell cultures. Strategies for defining pathways and uncovering rate-influencing steps in global metabolism, and applying this information for successful terpenoid metabolic engineering, are emphasized.     

Engineering Escherichia coli for production of functionalized terpenoids using plant P450s

Terpenoids are a highly diverse class of natural products that have historically provided a rich source for discovery of pharmacologically active small molecules, such as paclitaxel (Taxol) and artemisinin. Unfortunately, these secondary metabolites are typically produced in low abundance in their host organism, and their isolation consequently suffers from low yields and high consumption of natural resources. Furthermore, chemical synthesis of terpenoids can also be difficult to scale for industrial production. For these reasons, an attractive alternative strategy is to engineer metabolic pathways for production of pharmaceuticals or their precursors in a microbial host such as Escherichia coli. A key step is developing methods to carry out cytochrome P450 (P450)-based oxidation chemistry in vivo. Toward this goal, we have assembled two heterologous pathways for the biosynthesis of plant-derived terpenoid natural products, and we present the first examples of in vivo production of functionalized terpenoids in E. coli at high titer using native plant P450s.     

萜类生物合成的基因操作

萜类是一组结构迥异的化合物家族,其中很多具有较大的应用价值,如青蒿素和紫杉醇等,它们在多种微生物和植物中合成,但其天然产量低。萜类代谢工程通过DNA重组技术改造萜类合成细胞中的代谢途径,以提高萜类最终产量或在不含萜类的生物中合成萜类,为促进有用萜类合成提供了新的机会。以萜类化合物生物合成途径的基因转移与表达为切入点,综述了目前在微生物及植物中应用代谢工程提高萜类产量的研究进展。