放线菌天然产物糖基化改造研究进展

放线菌可以产生结构多样的天然产物, 其中包括很多重要的抗菌和抗肿瘤药物。糖基化修饰在天然产物中广泛存在, 糖基侧链的变化往往会影响天然产物的生物活性。本文综述了放线菌来源天然产物糖基化改造的研究进展。糖基侧链改造的方法主要分为体内基因工程和体外酶学法。运用这两种方法已经成功对多种天然产物进行了糖基侧链改造, 获得了大量带有新糖基修饰的天然产物, 其中有些生物活性得以提高。天然产物糖基侧链改造为新药开发提供了一个重要的途径。     

老兵又谱新传——新的糖基化方式、新的功能

在自然界中 ,糖类分子无所不在。虽然仍是那么一些单糖 ,但是在新的糖基转移酶作用下 ,以新的糖基化方式出现 ,就产生了新的功能。1.引子传统的概念是糖基转移酶定位在内质网和高尔基体中 ,它们的功能是合成糖苷键 ,在一组糖基转移酶的协同下 ,可以形成糖链。后来在哺乳动物精子的表面发现了半乳糖基转移酶 ,人们认识到糖基转移酶及其相关的糖链还可以参与细胞之间的识别和粘着。此后不仅在精子表面 ,而且在神经细胞表面也发现了一些糖基转移酶[1] 。在细胞质中发现Ｏ ＧｌｃＮＡｃ的糖基化及其对应的糖基转移酶和糖苷水解酶以后 ,极大地拓…     

糖核苷酸类化合物的合成与在生物学研究中的应用

糖基化是指在酶的作用下,蛋白质、脂质或小分子等连上糖类化合物的过程.它在生物体内广泛存在并有十分重要的生物学意艾.它是通过糖基转移酶催化完成的,其中糖核苷酸是糖基化过程中糖的供体.本文简要叙述了糖核苷酸类化合物的有机合成和生化合成的方法以及该类化合物在抗生素糖基化途径研究、糖基转移酶的生物化学研究和化学生物学机理研究中的应用.     

微生物合成黄酮糖苷类天然产物研究进展

黄酮糖苷类天然产物是植物中黄酮类化合物的主要存在形式,通过糖基化修饰,可以改变其水溶性、稳定性等,赋予其新的生物活性和功能。黄酮类化合物的糖基化修饰通常由植物源或微生物源的糖基转移酶催化,根据糖基的位置、类型和数量的不同,可形成多种类型的黄酮糖苷类产物。随着合成生物学和代谢工程的快速发展,在微生物中合成植物源黄酮糖苷类天然产物取得了重要进展。综述了糖基转移酶的聚类分析及糖基供体的途径改造,并对代谢工程优化黄酮糖苷类天然产物的微生物合成进行了分析讨论,并对其发展前景进行了展望。     

植物糖基转移酶的结构与机理及糖基化工程的研究进展

糖基转移酶(glycosyltransferases, GTs)广泛存在于各种有机体中,通过糖基化反应参与维持细胞代谢稳态.糖基转移酶能够识别多种受体,催化活化的糖基从供体分子转移到受体分子上,改变受体分子的化学稳定性、水溶性以及受体分子的转运能力和生物活性等,进而有助于提高其生物利用度和生物活性等.许多被糖基化修饰的化合物成为药物分子的重要来源.然而,天然产物中的糖苷类化合物存在含量低、提取难度大和提取产物纯度差等问题.在利用化学合成方法合成糖苷类化合物的过程中,无法实现特定位点的糖基化修饰,同时原料试剂和副产物易对环境造成污染.因此,近年来对糖基转移酶的研究日渐增多.本文简要综述了植物糖基转移酶的结构和生物技术应用的研究进展,为基于植物糖基转移酶结构的糖基化工程和生物活性糖苷化合物的生产提供有用信息.     

天然产物的C-糖基化研究进展

自然界中的天然产物在其结构上存在各种修饰,C-糖基化为其中一种比较稀少的修饰方式.C-C糖苷键的形成由C-糖基转移酶负责催化.含C-糖基的化合物多数来自微生物,但高等植物也会有少量积累.综述了近些年来在天然产物C-糖基化方面的研究工作,并对其在药物开发方面的潜力进行了展望.     

植物源UDP-糖基转移酶及其分子改造*

糖基化能够增加化合物的结构多样性,有效改善水溶性、药理活性和生物利用度,对植物天然产物的药物开发至关重要。UDP-糖基转移酶(UGTs)能够催化糖基从活化的核苷酸糖供体转移到受体形成糖苷键,植物中天然产物的糖基化修饰主要通过UGTs实现。但是大多数天然UGTs的催化活性、稳定性和底物特异性较低,难以满足工业用酶的要求,限制了它们的工业化应用。近年来,通过分子改造技术改进天然UGTs的催化特性取得了突破性的研究进展。为此,概述了植物源UGTs的挖掘与表征、三维结构和催化机制,归纳了UGTs分子改造的思路和方法,包括理性设计和定向进化,重点介绍了结构域替换、序列保守分析和结构分析与定点突变的结合,总结了定向进化中的高通量筛选方法,为植物天然产物酶法糖基化的工业应用提供了参考和借鉴。     

酵母表达人源化糖蛋白研究进展

与人体天然复杂型糖蛋白相比,使用酵母生产的药用蛋白带有高甘露糖型N-糖链。这一差异在临床应用中产生了许多不良影响。目前,可以通过消除酵母特有的内源糖基化反应,引入哺乳动物细胞中的一系列糖基转移酶及转运蛋白对酵母糖基化路径进行改造,从而使其表达出人源化的复杂型N-聚糖。本文介绍了酵母N-糖基化特点、糖基化不均一性,综述了近年来利用基因工程改造酵母N-糖基化路径获得特定的人源N-连接糖蛋白以及使用内切糖苷酶生产人源糖蛋白的研究进展,并且对存在的问题及今后的发展前景进行了讨论。     

分子伴侣协助下抗肿瘤抗生素美达霉素生物合成中的糖基转移酶Med-ORF8 的原核表达

糖基转移酶在天然产物的糖基化修饰过程中起关键作用。目前链霉菌源抗肿瘤抗生素美达霉素(Medermycin)生物合成途径中的糖基转移酶Med-ORF8的原核表达及酶学性质还未有研究。首先通过结构模拟确定Med-ORF8的端部加上His-标签不会影响其三维结构的正确折叠,然后利用2种pET原核表达载体来进行Med-ORF8的原核表达,发现以pET-28a(+)为载体进行表达时,目的蛋白产量非常高,但是以不可溶的包涵体形式为主。当分子伴侣基因(编码大肠杆菌触发因子)与Med-ORF8的编码基因共表达时,在优化诱导条件的情况下,可以有效减少包涵体的形成,提高了Med-ORF8的可溶性表达效率,为Med-ORF8的酶学分析打下基础。     

糖基转移酶和去糖基化酶

在糖基化工程中,通过酶法对蛋白质进行糖基化修饰和对天然糖蛋白去糖基化是研究糖蛋白结构与功能的重要手段。本文综述了近年来所纯化的主要的糖基化转移酶和去糖基化酶的性质和应用。     

Features and applications of bacterial glycosyltransferases: current state and prospects

The bioactivity of many natural products including valuable antibiotics and anticancer therapeutics depends on their sugar moieties. Changes in the structures of these sugars can deeply influence the biological activity, specificity and pharmacological properties of the parent compounds. The chemical synthesis of such sugar ligands is exceedingly difficult to carry out and therefore impractical to establish on a large scale. Therefore, glycosyltransferases are essential tools for chemoenzymatic and in vivo approaches for the development of complex glycosylated natural products. In the last 10 years, several examples of successful alteration and diversification of natural product glycosylation patterns via metabolic pathway engineering and enzymatic glycodiversification have been described. Due to the relaxed substrate specificity of many sugar biosynthetic enzymes and glycosyltransferases involved in natural product biosynthesis, it is possible to obtain novel glycosylated compounds using different methods. In this review, we would like to provide an overview of recent advances in diversification of the glycosylated natural products and glycosyltransferase engineering.     

Achievements and impacts of glycosylation reactions involved in natural product biosynthesis in prokaryotes

Bioactive natural products, such as polyketides, flavonoids, glycopeptides, and aminoglycosides, have been used as therapeutic agents. Many of them contain structurally diverse sugar moieties attached to the aglycone core structures. Glycosyltransferases (GTs) catalyze the attachment of nucleotide-activated sugar substrates to acceptor aglycones. Because these sugar moieties are usually essential for biological activity, in vivo pathway engineering in prokaryotic hosts and in vitro enzymatic approaches coupled with GT engineering are currently being used to synthesize novel glycosylated derivatives, and some of them exhibited improved biological activities compared to the parent molecules. Therefore, harnessing the potential of diverse glycosylation reactions in prokaryotes will increase the structural diversity of natural products and the possibility to generate new bioactive products.     

Engineering the glycosylation of natural products in actinomycetes

Bioactive natural products are frequently glycosylated with saccharide chains of different length, in which the sugars contribute to specific interactions with the biological target. Combinatorial biosynthesis approaches are being used in antibiotic-producing actinomycetes to generate derivatives with novel sugars in their architecture. Recent advances in this area indicate that glycosyltransferases involved in the biosynthesis of natural products have substrate flexibility regarding the sugar donor but also, less frequently, with respect to the aglycon acceptor. Therefore, the possibility exists of altering the glycosylation pattern of natural products, thus enabling an increase in the structural diversity of natural products.     

Enzymatic tools for engineering natural product glycosylation

Glycosylated natural products have served as reliable platforms for the development of many existing front-line drugs. In an effort to explore the contribution of the sugar constituents of these compounds, research groups have focused upon the development of chemical and enzymatic tools to diversify natural product glycosylation. Among the complementary routes available, in vivo pathway engineering, also referred to as 'combinatorial biosynthesis', is an emerging method that relies upon the co-expression of sugar biosynthetic gene cassettes and glycosyltransferases in a host organism to generate novel glycosylated natural products. An overview of recent progress in combinatorial biosynthesis is highlighted in this review, emphasizing the elucidation of nucleotide-sugar biosynthetic pathways and recent developments on glycosyltransferases.     

Metabolic and functional diversity of saponins,biosynthetic intermediates and semi-synthetic derivatives

Abstract

Saponins are widely distributed plant natural products with vast structural and functional diversity. They are typically composed of a hydrophobic aglycone, which is extensively decorated with functional groups prior to the addition of hydrophilic sugar moieties, to result in surface-active amphipathic compounds. The saponins are broadly classified as triterpenoids, steroids or steroidal glycoalkaloids, based on the aglycone structure from which they are derived. The saponins and their biosynthetic intermediates display a variety of biological activities of interest to the pharmaceutical, cosmetic and food sectors. Although their relevance in industrial applications has long been recognized, their role in plants is underexplored. Recent research on modulating native pathway flux in saponin biosynthesis has demonstrated the roles of saponins and their biosynthetic intermediates in plant growth and development. Here, we review the literature on the effects of these molecules on plant physiology, which collectively implicate them in plant primary processes. The industrial uses and potential of saponins are discussed with respect to structure and activity, highlighting the undoubted value of these molecules as therapeutics.     

Glycosyltransferases involved in the biosynthesis of biologically active natural products that contain oligosaccharides

A unique characteristic of carbohydrates is their structural diversity which is greater than that of many other classes of biological compounds. Carbohydrate-containing natural products show many different biological activities and some of them have been developed as drugs for medical use. The biosynthesis of carbohydrate-containing natural products is catalysed by glycosyltransferases. In this review we will present information on the function of glycosyltransferases involved in the biosynthesis of oligosaccharide antibiotics focusing especially on urdamycins and landomycins, two angucycline antibiotics with interesting antitumor activities. We will also discuss the use of glycosyltransferases in combinatorial biosynthesis to generate new "hybrid" antibiotics.     

Mechanisms and structures of vitamin B(6)-dependent enzymes involved in deoxy sugar biosynthesis

PLP is well-regarded for its role as a coenzyme in a number of diverse enzymatic reactions. Transamination, deoxygenation, and aldol reactions mediated by PLP-dependent enzymes enliven and enrich deoxy sugar biosynthesis, endowing these compounds with unique structures and contributing to their roles as determinants of biological activity in many natural products. The importance of deoxy aminosugars in natural product biosynthesis has spurred several recent structural investigations of sugar aminotransferases. The structure of a PMP-dependent enzyme catalyzing the C-3 deoxygenation reaction in the biosynthesis of ascarylose was also determined. These studies, and the crystal structures they have provided, offer a wealth of new insights regarding the enzymology of PLP/PMP-dependent enzymes in deoxy sugar biosynthesis. In this review, we consider these recent achievements in the structural biology of deoxy sugar biosynthetic enzymes and the important implications they hold for understanding enzyme catalysis and natural product biosynthesis in general. This article is part of a Special Issue entitled: Pyridoxal Phosphate Enzymology.     

Computational-aided design of a library of lactams through a diversity-oriented synthesis strategy

Small molecule libraries for virtual screening are becoming a well-established tool for the identification of new hit compounds. As for experimental assays, the library quality, defined in terms of structural complexity and diversity, is crucial to increase the chance of a successful outcome in the screening campaign. In this context, Diversity-Oriented Synthesis has proven to be very effective, as the compounds generated are structurally complex and differ not only for the appendages, but also for the molecular scaffold. In this work, we automated the design of a library of lactams by applying a Diversity-Oriented Synthesis strategy called Build/Couple/Pair. We evaluated the novelty and diversity of these compounds by comparing them with lactam moieties contained in approved drugs, natural products, and bioactive compounds from ChEMBL. Finally, depending on their scaffold we classified them into β-, γ-, δ-, ε-, and isolated, fused, bridged and spirolactam groups and we assessed their drug-like and lead-like properties, thus providing the value of this novel in silico designed library for medicinal chemistry applications.     

Structure, mechanism and engineering of plant natural product glycosyltransferases

Glycosylation is a key mechanism in determining chemical complexity and diversity of plant natural products, and influencing their chemical properties and bioactivities. Uridine diphosphate glycosyltransferases (UGTs) are the central players in these glycosylation processes for decorating natural products with sugars. Crystal structures of plant UGTs have revealed their exquisite architectures and provided the structural basis for understanding their catalytic mechanism and substrate specificity. Structure-based UGT engineering can alter substrate specificity; compromise or enhance catalytic efficiency; and confer reversibility to the glycosylation reaction. This review highlights the structural insights on plant UGTs and successes in glycosylation engineering.     

Chemoenzymatic and template-directed synthesis of bioactive macrocyclic peptides.

Non-ribosomally synthesized peptides have compelling biological activities ranging from antimicrobial to immunosuppressive and from cytostatic to antitumor. The broad spectrum of applications in modern medicine is reflected in the great structural diversity of these natural products. They contain unique building blocks, such as d-amino acids, fatty acids, sugar moieties, and heterocyclic elements, as well as halogenated, methylated, and formylated residues. In the past decades, significant progress has been made toward the understanding of the biosynthesis of these secondary metabolites by nonribosomal peptide synthetases (NRPSs) and their associated tailoring enzymes. Guided by this knowledge, researchers genetically redesigned the NRPS template to synthesize new peptide products. Moreover, chemoenzymatic strategies were developed to rationally engineer nonribosomal peptides products in order to increase or alter their bioactivities. Specifically, chemical synthesis combined with peptide cyclization mediated by nonribosomal thioesterase domains enabled the synthesis of glycosylated cyclopeptides, inhibitors of integrin receptors, peptide/polyketide hybrids, lipopeptide antibiotics, and streptogramin B antibiotics. In addition to the synthetic potential of these cyclization catalysts, which is the main focus of this review, different enzymes for tailoring of peptide scaffolds as well as the manipulation of carrier proteins with reporter-labeled coenzyme A analogs are discussed.