有机化合物生物催化研究进展

随着近十几年来工业生物技术的发展,有机化合物的生物催化也取得了飞速的进步.近几年的研究集中在:新生物催化剂的筛选和酶的定向改造;非水相生物催化中酶有机溶剂耐受性的增强和非传统介质的应用;生物催化在手性化合物,药物等精细化学品领域的应用;组合生物催化作为组合化学和生物催化相结合而成的一个新技术生长点,并取得一定的进展,为新药的开发提供一种切实可行的方法.     

RNA的生物催化功能及其研究进展

长期以来,我们一直认为生命过程中的所有化学反应是在生物催化剂——酶的作用下进行的,而所有的酶都是蛋白质或带有辅基的蛋白质。但近年研究发现,某些RNA分子也具有“酶”的生物催化功能,它们能在一定条件下催化自身或其它RNA分子发生化学反应。Cech(1981,1986)据此提出了RNA生物催化剂的概念,将这些具有酶活性的RNA称之为核酶(ribozyme)。本文简要介绍几种RNA分子的生物催化功能及其研究进展。     

催化混杂性驱动的酶功能重塑

作为生物催化剂,酶蛋白介导的生化反应具有条件温和、绿色环保等优点。然而相比化学催化剂,天然酶功能的局限性制约了它在生物制造领域的广泛应用。前期研究表明,酶蛋白除了催化专一性外,同时还展现出混杂性的一面,可在特定条件下催化非天然模式反应。这一特性为酶分子功能重塑提供了新思路,可用来指导人工酶设计,拓展天然酶的催化边界,实现新颖酶促反应类型,以扩大酶催化应用场景。本文从酶催化功能混杂性背后可能的进化机制入手,综述了当前诱导酶催化功能混杂性的常用策略,如定向进化、构象动力学、反应条件诱导及祖先酶重构等技术,并从催化机制、构效关系及适应性进化等多个角度,结合近年来相关研究实例,探讨了催化功能混杂性背后的分子机制,为突破天然酶促反应局限性、创制催化非天然反应的高效人工酶元件提供参考。     

微囊载体固定化多酶的研究进展

多酶催化是利用多种生物酶构建反应体系或网络,在生物体外实现化学品的合成.在生物制造过程中,多酶的共固定化有利于提高酶的稳定性和重复使用率,更利于多酶间的协同催化.在精准调控下,多酶固定化载体的微囊材料有望实现多酶协同催化性能的最大化.本文中,笔者分析了微囊体系的特点,综述了微囊材料及其固定化多酶的优缺点,总结了微囊多酶...     

多酶共固定化反应体系的研究进展

近年来,生物催化为化学、生物学和生物工程学等领域提供了一种绿色研究工具,其中多酶体系在这些领域中的应用越来越受到关注,其克服了以往单个酶不能满足催化需求的局限性,同时多酶共固定化在级联反应过程中,可增加酶周围的反应物浓度,并将不同酶的催化特性结合起来,能排除干扰因素,从而提高酶的整体催化效率。对多酶共固定化反应体系的研究进展进行了综述,包括多酶反应体系的类别、共固定化技术的特点以及相关应用,并对共固定化多酶反应体系进行了展望。     

工业蛋白质理性设计与应用

作为合成生物学与绿色生物制造等领域的底层核心技术,蛋白理性设计可有效解决天然功能元件性能不足等共性挑战,创制高性能人工酶元件。值此天津工业生物研究所(Tianjin Institute of Industrial Biotechnology, TIB)创立10周年之际,文中回顾了研究所在工业蛋白理性设计领域的系列重要工作进展。从酶设计方法学研究、新酶反应设计到生物催化应用等方面进行了分析讨论,并展望了本领域未来发展方向。望借此搭建学术界和产业界与酶理性设计的桥梁,促进新技术、新策略的开发应用,加速融合人工酶的基础研究与产业应用,推动我国生物制造领域的科技创新升级。     

微生物途径发展生物柴油产业

生物柴油实际上就是生物油脂与甲醇或乙醇在酸、碱催化剂的作用下进行脂交换反应而制造的脂肪酸甲酯或乙酯;也可以在常温下由微生物脂酶催化进行酯化反应,其产品是一种可再生燃料,能替代石油柴油。这些生物柴油主要来自植物油或其它生物油脂,也有用废弃食用油为原料通过甲醇的酯交换反应来制造生物柴油的。研发这些生物柴油也可以说是节能的一项重要措施。在我国,对石油的需求量越来越大,石油进口量也随之猛增,显示出我国的能源形势日益严峻。面对这种情况,发展可再生能源或替代能源是个必然趋势,生物柴油便是其中之一。目前我国生物柴油的…     

反应温度调控生物催化立体选择性的研究进展

立体选择性调控是手性生物催化的重要内容。反应温度调节作为一种简单、便捷的立体选择性调控方式已在生物催化制备手性化学品中发挥了重要作用。分析了温度引起酶立体选择性变化的机理,并综述了该方法在脂肪酶、酯酶、醇脱氢酶、青霉素G酰化酶和脱卤酶等酶催化反应中的应用。     

工业生物催化前线动态及名家观点

随着现代生物技术的进步,尤其是酶的快速筛选和活力优化技术的发展,使酶的获取更加容易、酶的操作更加简单,进而促使生物催化成为手性合成的便利工具。综述了一些著名的国际化工或制药公司最近在生物催化技术研发和应用方面的动态信息以及相关技术的一些评论,以便我国从事工业生物催化工作的相关人士能从中获得有益启示。     

脂肪酶催化合成生物柴油的瓶颈问题及其对策研究进展

生物柴油,一种新型的清洁能源燃料,具有可再生、可生物降解、环境友好等优良的品性,可部分或全部替代石化柴油。碱催化法、脂肪酶催化法及超临界法是合成生物柴油的主要工艺,其中脂肪酶催化法是一种节能型、环保型工艺,在节能和环保方面,有着碱催化法无可比拟的优越性,具有良好的工业应用前景。但目前在实现产业化的进程中仍存在如酶成本高、稳定性较差、甲醇对酶的失活效应及反应时间长等瓶颈问题。通过固定化技术和全细胞催化剂的采用、甲醇流加方式的改进、溶剂工程的改善及酰基受体和耐醇酶的开发等技术手段,结合固定床生物反应器,较好地解决了这些瓶颈问题,从而推进了酶催化法合成生物柴油的工业化进程。本文主要对酶法合成生物柴油工艺存在的主要问题及相应对策研究进展进行概括介绍,并对其工业化发展前景进行讨论。     

Combinatorial biocatalysis: taking the lead from nature

Combinatorial biocatalysis is an emerging technology in the field of drug discovery. The biocatalytic approach to combinatorial chemistry uses enzymatic, chemoenzymatic, and microbial transformations to generate libraries from lead compounds. Important recent advances in combinatorial biocatalysis include iterative derivatization of small molecules and complex natural products, regioselectively controlled libraries, novel one-pot library syntheses, process automation, and biocatalyst enhancements.     

Biocatalysis in natural products chemistry

The properties of enzymes and microbial cells as biocatalysts useful in natural products chemistry are discussed from the perspective of the chemical transformations they catalyse. Attention is focused on numerous reactions of value to natural products chemists, including the acyloin condensation, Baeyer-Villiger oxidation, regio- and enantioselective ester hydrolyses, oxidations of aromatic and non-aromatic substrates, oxidoreduction and O- and N-dealkylations. Compounds considered in this review include amino acids, alkaloids, antibiotics, coumarins, naphthoquinones, quassinoids, rotenoids and mono-, sesqui-, di- and triterpenoid substrates. The value of biocatalysis compared with traditional chemical catalysis is considered within the broad framework of natural products chemistry, and the potential for using immobilized enzyme and cell technology is presented.     

Biocatalytic synthesis of hydroxylated natural products using aldolases and related enzymes

Synthetic building blocks bearing hydroxylated chiral centers are important targets for biocatalysis. Many C-C bond forming enzymes have recently been investigated for new applications and new strategies towards the synthesis of natural products and related oxygenated compounds. Several old catalysts have been studied to increase our functional knowledge of natural aldolase-type enzymes, and new mutated catalysts or catalytic antibodies have been tested for their synthetic utility.     

生物信息学在工业生物催化研究中的应用

随着石油等不可再生资源的日益减少以及环境污染问题的日益严重,应用工业生物催化技术改造或取代传统化工工艺已经成为新世纪化学工业可持续发展的研究热点。工业生物催化技术的研究对象是生物催化剂及其催化过程。近来,利用生物信息学技术进行工业生物催化研究已经越来越受到人们的重视。随着工业生物催化的发展,生物信息学将直接指导并加快新型高效生物催化剂的发现及功能改造进程。     

Application of combinatorial biocatalysis for a unique ring expansion of dihydroxymethylzearalenone

Combinatorial biocatalysis was applied to generate a diverse set of dihydroxymethylzearalenone analogs with modified ring structure. In one representative chemoenzymatic reaction sequence, dihydroxymethylzearalenone was first subjected to a unique enzyme-catalyzed oxidative ring opening reaction that creates two new carboxylic groups on the molecule. These groups served as reaction sites for further derivatization involving biocatalytic ring closure reactions with structurally diverse bifunctional reagents, including different diols and diamines. As a result, a library of cyclic bislactones and bislactams was created, with modified ring structures covering chemical space and structure activity relationships unattainable by conventional synthetic means.     

手性技术与生物催化

简要介绍了手性,手性技术与生物催化的基本概念。手性,是指一个有机分子具有不对称性,形成两种空间排布方式不同的对映异构体。手性技术即生产手性化合物的技术,手性化合物的制备方法主要有手性源、外消旋体拆分、不对称合成等几种。生物催化,即利用酶或微生物等生物材料催化进行某种化学反应,被认为是手性化合物生产取得突破的关健技术。文章还介绍了生物催化外消旋体拆分、生物催化不对称合成等几种生产手性化合物的应用实例。     

Accelerating whole-cell biocatalysis by reducing outer membrane permeability barrier

Whole-cell biocatalysts are preferred in many biocatalysis applications. However, due to permeability barriers imposed by cell envelopes, whole-cell catalyzed reactions are reportedly 10-100-fold slower than reactions catalyzed by free enzymes. In this study, we accelerated whole-cell biocatalysis by reducing the membrane permeability barrier using molecular engineering approaches. Escherichia coli cells with genetically altered outer membrane structures were used. Specifically, a lipopolysaccarides mutant SM101 and a Braun's lipoprotein mutant E609L were used along with two model substrates that differ substantially in size and hydrophobicity, nitrocefin, and a tetrapeptide N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide. The reduction of the outer membrane permeability by genetic methods led to significant increases (up to 380%) in reaction rates of whole-cell catalyzed reactions. The magnitude of increase in biocatalysis rates was dependent on the substrates and on the nature of mutations introduced in the outer membrane structure. Notably, mutations in outer membrane can render the outer membrane completely permeable to one substrate, a barrierless condition that maximizes the reaction rate. The impact of the mutations introduced on the permeability barrier of the membranes was compared to the impact of polymixin B nonapeptide, a known potent permeabilizer acting on lipopolysaccharides. Our results suggest that genetic modifications to enhance the permeability of hydrophilic molecules should target the Lipid A region. However, strategies other than reduction of Lipid A synthesis should be considered. As we have demonstrated with tetrapeptide, membrane engineering can be much more effective in reducing a permeability barrier than are exogenous permeabilizers. This work, to our knowledge, is the first use of a molecular membrane engineering approach to address substrate permeability limitations encountered in biocatalysis applications.     

Homogeneous Biocatalysis in Organic Solvents and Water-Organic Mixtures

Biocatalysis in non-aqueous media has undergone tremendous development during the last decade, and numerous reactions have been introduced and optimized for synthetic applications. In contrast to aqueous enzymology, biotransformations in organic solvents offer unique industrially attractive advantages, such as: drastic changes in the enantioselectivity of the reaction, the reversal of the thermodynamic equilibrium of hydrolysis reactions, suppression of water-dependent side reactions, and resistance to bacterial contamination. Currently, the field is dominated by heterogeneous biocatalysis based primarily on lyophilized enzyme powders, cross-linked crystals, and enzymes immobilized on inert supports that are mainly applied in enantioselective synthesis. However, low reaction rates are an inherent problem of the heterogeneous biocatalysis, while the homogeneous systems have the advantage that the elimination of diffusional barriers of substrates and products between organic and water phases results in an increase in the reaction rate. Here the discussion is focused on the correlation between activity and structure of the intact enzymes dissolved in neat organic solvents, as well as modifications of natural enzymes, which make them soluble and catalytically active in non-aqueous environment. Factors that influence conformation and stability of the enzymes are also discussed. Current developments in non-aqueous biocatalysts that combine advantages of protein modification and immobilization, i.e., HIP plastics, enzyme chips, ionic liquids, are introduced. Finally, engineering enzymes for biotransformations in non-conventional media by directed evolution is summarized.     

聚酮类抗生素组合生物合成

组合生物合成是公认的产生大量"非天然"的天然产物的一种有效方法,也是近年来药物创新与应用的研究热点和重要手段之一。目前,组合生物合成在聚酮类抗生素等生物活性物质的开发应用研究中已经取得了显著的成果。结合文献中的例子,回顾了运用组合生物合成在天然产物的基础上产生更多结构及功能多样性的聚酮类抗生素的方法和思路,并对某些方法所存在的问题与不足进行了讨论。