微生物发酵法生产S-腺苷甲硫氨酸的研究进展

S-腺苷甲硫氨酸(S-adenosyl-l-methionine, SAM)广泛存在于生物体内,主要参与生物体内的转甲基过程、转硫过程及转氨丙基过程,具有重要的生理功能,其生产备受重视。目前SAM生产的研究主要集中于微生物发酵法,该方法与化学合成法和酶催化法相比,成本较低且更容易实现工业化生产。随着需求量的迅速增加,通过菌种改良提高SAM产量备受关注。当前SAM生产菌种改良的主要策略包括常规育种和代谢工程。本文综述了提高微生物生产SAM能力的近期研究进展并探讨了SAM生产中的瓶颈问题及解决方法,以期为进一步提高SAM产量提供思路。     

微生物转化生产S-腺苷甲硫氨酸

S-腺苷甲硫氨酸(S-adenosyl-L-methionine,SAM)是具有广阔市场前景的活性氨基酸,微生物转化法是近年来报道较多的SAM生产方法.综合近年来SAM生产菌株的基因改造和发酵优化方面的进展,从提高SAM合成酶表达和酶活、优化甲醇和甘油的流加方式、改善ATP的生成和L-甲硫氨酸的补料、阻断下游代谢路径等方面,综述了促进SAM合成及其积累的多重策略及机制.最后结合笔者多年研究实践,讨论了微生物转化生产SAM的未来研究方向.     

色氨酸发酵的研究

<正> L-色氨酸是人和动物体内必需氨基酸之一,在医药、食品和饲料添加方面有着广泛的用途。由于它在动植物蛋白中含量很低,从天然蛋白质中提取来源有限,而化学合成法又代价较高,因此用发酵法来生产L-色氨酸长期以来受到重视。目前主要有直接发酵法,前体发酵法和酶促转化法三种,据报导,1973年Snilo等人用B·subtilis FT~r直接发酵生产L色氨酸,产量达6.1克/升。1973年Nagino等人以及1974、1975年Nagino和NaRaymu等人成功地诱导各种     

多胺生物合成途径中两个关键酶基因研究进展

多胺是一类小分子生物活性物质,广泛存在于生物体内,与植物的生长发育、衰老及抗逆性都有着密切的联系。就多胺合成途径中的两个关键酶基因,即S-腺苷甲硫氨酸合成酶基因(SAMS)和S-腺苷甲硫氨酸脱羧酶基因(SAMDC)的克隆、表达,以及转S-腺苷甲硫氨酸合成酶基因(SAMS)和转S-腺苷甲硫氨酸脱羧酶基因(SAMDC)表达调控等方面的研究进行回顾总结,并对其应用前景进行展望。     

用微生物酶合成旋光的硫和硒氨基酸

<正> 自然界中存在各种硫化合物,而且大多数是生理活性的。甲硫氨酸、半胱氨酸和其它一些硫氨基酸尤其起着重要的代谢作用。例如,L-甲硫氨酸不仅是一种必需氨基酸,而且是通过中间体S-腺苷-L-甲硫氨酸的各种转甲基酶系中的主要供体。它还是多胺和一种植物激素—乙烯的重要前体。大量的DL-甲硫氨酸广泛用作黄豆粉和其它缺乏硫氨基酸的饲料的添加剂。甲硫氨酸、半胱氨酸和其它氨基酸还是药物、化妆品和化学品的很重要的原料。目前利用细菌可以由简单的碳、氨源生产各种L-氨基酸,包括谷氨酸和赖氨酸。但用微生物方法生产硫氨基酸则很少获得成功。     

长链二元酸发酵菌种创制和工艺研究进展

长链二元酸作为合成多种高附加值化学品的原料,已广泛应用于化工、农业和医药等领域,目前全球对于长链二元酸的需求呈逐年增长态势。化学法合成长链二元酸对反应条件要求严苛且工艺复杂,而微生物发酵合成在经济性和难易度等方面具有无可比拟的优势。本文综述了长链二元酸的合成方法,包括化学合成法和微生物发酵法,分子工程选育高产菌株的进展以及生物发酵法生产长链二元酸的产业化现状,并就其存在的问题进行了探讨,最后对合成生物学创制长链二元酸高产菌株进行了总结和展望。     

腺苷甲硫氨酸合成酶的基因及结构研究进展

腺苷甲硫氨酸合成酶催化ATP和L-甲硫氨酸合成腺苷甲硫氨酸,在不同生物体和不同组织中腺苷甲硫氨酸合成酶的存在形式和编码酶的基因都有差别,本文综述了不同生物的腺苷甲硫氨酸合成酶的基因、酶结构、酶反应动力学及应用前景。     

离子交换吸附L-色氨酸的研究

L-色氨酸是含吲哚环的芳香族氨基酸,也是人类必需的氨基酸之一,目前,工业上除部分采用化学合成法外,主要采用前体发酵法和酶促转化法生产。随着在医药、食品和饲料加工中应用的开拓,市场的需求量日益增加,色氨酸生产具有广阔的前途。     

微生物合成腺苷蛋氨酸的研究进展

腺苷蛋氨酸(SAM)是一种存在于所有细胞中的重要代谢产物,对肝病、抑郁症等疾病具有良好疗效,并且无任何副作用,市场需求巨大。目前,国内外学者对SAM的生物合成进行了广泛的研究,一方面在传统发酵法工业化生产SAM的基础上进一步改进发酵策略;另一方面通过代谢工程优化宿主菌代谢网络或者利用基因电路等巧妙筛选方法来进行理性化育种。综述了微生物高效生产SAM的研究进展,讨论了微生物高效生产SAM的意义及存在的问题,旨为提高SAM工业化生产效率提供指导性意义。     

腺苷甲硫氨酸合成酶的提取纯化研究

腺苷蛋氨酸具有转甲基、转硫和转氨丙基等重要生理作用,已成为治疗疾病的重要药物。目的：为腺苷蛋氨酸合酶的基因克隆做准备。方法：研究了腺苷甲硫氨酸合成酶的提取和纯化。腺苷蛋氨酸合酶为胞内酶,其提取需先进行细胞破碎,然后进行盐析和离子交换层析等方法来纯化。酵母的破壁试验考察了研磨、加入有机溶剂和超声波等不同的破碎方法。结果：超声波破碎法最好,得到粗酶液的酶活力为0．934U／ml;经过硫酸铵盐析后,利用离子交换层析法纯化腺苷甲硫氨酸合成酶,作出了腺苷甲硫氨酸合成酶的穿透曲线和洗脱曲线。     

生物技术生产丁酸的研究进展

丁酸作为一种重要的化工原料,已经广泛应用于食品添加剂与医药等领域。目前,工业上生产丁酸主要是从石油中提取有机化合物进行化学合成。与有机化合物合成法相比,微生物发酵产丁酸的优势有：所用的原料来源非常广,发酵过程低能耗,不污染环境,而且可以持续添加原料发酵生产丁酸。因此,通过生物技术发酵生产丁酸越来越受到人们的重视。介绍了丁酸的性质、产丁酸菌株的特点、微生物发酵产丁酸的细胞代谢途径及其调控、发酵法生产丁酸的工艺运行方式和产丁酸菌株及其代谢产物的生理功能这五部分内容,以期为今后开展发酵法产丁酸的微生物基因工程改造以及生产工艺的优化提供参考。     

Progress in the research of S-adenosyl-l-methionine production

This minireview mainly aims at the study of S-adenosyl-l-methionine (SAM) production by microbial fermentation. A brief introduction of the biological role and application of SAM was presented. In general, SAM production can be improved by breeding of the producing strain through the conventional mutation or genetic engineering approach in the molecular or cellular scale, by optimization of culture conditions in the cellular scale or bioreactor engineering scale, or by multiscale approach. The productivity of SAM fermentation has been improved greatly through the efforts of many researchers using the methods previously mentioned. The SAM-producing strains used extensively are Pichia pastoris and Saccharomyces cerevisiae. The effect of SAM on antibiotic production was also exemplified. The skill and scheme beneficial to the improvement of SAM production involves the enhancement of SAM synthetase (methionine adenosyltransferase) activity and selection of engineered constitutive promoters with appropriate strength; seeking for and eliminating the rate-limiting factors in SAM synthesis, namely, knocking off the genes that transform SAM and l-methionine (L-Met) to cysteine; release the feedback inhibition of SAM to methylenetetrahydrofolate reductase; blocking the transsulfuration pathway by interfering the responsible enzymes; enhancing ATP level through pulsed feeding of glycerol; and optimizing the L-Met feeding strategy. Precise control of gene expression and quantitative assessment of physiological parameters in engineered P. pastoris were highlighted. Finally, a discussion of the prospect of SAM production was presented.     

Microbial production of metabolites and associated enzymatic reactions under high pressure

High environmental pressure exerts an external stress on the survival of microorganisms that are commonly found under normal pressure. In response, many growth traits alter, including cell morphology and physiology, cellular structure, metabolism, physical and chemical properties, the reproductive process, and defense mechanisms. The high-pressure technology (HP) has been industrially utilized in pressurized sterilization, synthesis of stress-induced products, and microbial/enzymatic transformation of chemicals. This article reviews current research on pressure-induced production of metabolites in normal-pressure microbes and their enzymatic reactions. Factors that affect the production of such metabolites are summarized, as well as the effect of pressure on the performance of microbial fermentation and the yield of flavoring compounds, different categories of induced enzymatic reactions and their characteristics in the supercritical carbon dioxide fluid, effects on enzyme activity, and the selection of desirable bacterial strains. Technological challenges are discussed, and future research directions are proposed. Information presented here will benefit the research, development, and application of the HP technology to improve microbial fermentation and enzymatic production of biologically active substances, thereby help to meet their increasing demand from the ever-expanding market.     

微生物发酵法生产L-色氨酸的代谢工程研究

L-色氨酸作为人体内的一种必需氨基酸,广泛应用于医药、食品与饲料等行业.工业上采用的色氨酸生产方法有化学合成法、转化法及微生物发酵法.近年来,随着代谢工程在色氨酸菌种选育中的成功运用,微生物发酵法逐渐成为主要的色氨酸生产方法.系统综述了微生物发酵法生产色氨酸所涉及的代谢工程策略,包括生物合成色氨酸的代谢调控机制以及途径...     

Recent advances in producing food additive L-malate: Chassis,substrate, pathway,fermentation regulation and application

In addition to being an important intermediate in the TCA cycle, L-malate is also widely used in the chemical and beverage industries. Due to the resulting high demand, numerous studies investigated chemical methods to synthesize L-malate from petrochemical resources, but such approaches are hampered by complex downstream processing and environmental pollution. Accordingly, there is an urgent need to develop microbial methods for environmentally-friendly and economical L-malate biosynthesis. The rapid progress and understanding of DNA manipulation, cell physiology, and cell metabolism can improve industrial L-malate biosynthesis by applying intelligent biochemical strategies and advanced synthetic biology tools. In this paper, we mainly focused on biotechnological approaches for enhancing L-malate synthesis, encompassing the microbial chassis, substrate utilization, synthesis pathway, fermentation regulation, and industrial application. This review emphasizes the application of novel metabolic engineering strategies and synthetic biology tools combined with a deep understanding of microbial physiology to improve industrial L-malate biosynthesis in the future.     

Progress in the microbial production of <Emphasis Type="Italic">S</Emphasis>-adenosyl-<Emphasis Type="SmallCaps">l</Emphasis>-methionine

S-Adenosyl-l-methionine (SAM), which exists in all living organisms, serves as an activated group donor in a range of metabolic reactions, including trans-methylation, trans-sulfuration and trans-propylamine. Compared with its chemical synthesis and enzyme catalysis production, the microbial production of SAM is feasible for industrial applications. The current clinical demand for SAM is constantly increasing. Therefore, vast interest exists in engineering the SAM metabolism in cells for increasing product titers. Here, we provided an overview of updates on SAM microbial productivity improvements with an emphasis on various strategies that have been used to enhance SAM production based on increasing the precursor and co-factor availabilities in microbes. These strategies included the sections of SAM-producing microbes and their mutant screening, optimization of the fermentation process, and the metabolic engineering. The SAM-producing strains that were used extensively were Saccharomyces cerevisiae, Pichia pastoris, Candida utilis, Scheffersomyces stipitis, Kluyveromyces lactis, Kluyveromyces marxianus, Corynebacterium glutamicum, and Escherichia coli, in addition to others. The optimization of the fermentation process mainly focused on the enhancement of the methionine, ATP, and other co-factor levels through pulsed feeding as well as the optimization of nitrogen and carbon sources. Various metabolic engineering strategies using precise control of gene expression in engineered strains were also highlighted in the present review. In addition, some prospects on SAM microbial production were discussed.     

Levan果聚糖的应用与生产研究进展

Levan是一类果聚糖,由大量的果糖单元以β-（2,6）果糖苷键连接构成聚糖主链并含有少量β-（2,1）果糖苷键连接的支链组成。部分微生物来源的Levan具有抗肿瘤、抗病毒、降血糖、降血脂、免疫增强等重要的生物活性,在医药和功能性食品方面具有巨大的应用潜能。微生物发酵液提取和酶法合成是目前大量获得Levan果聚糖的两种方法,其中微生物发酵液提取的Levan果聚糖产量和蔗糖转化率一般较低,且发酵液中同时存在的其他高聚物不利于Levan的规模化纯化;而利用Levan蔗糖酶以蔗糖为底物转果糖基合成的Levan果聚糖产量已经高达200g/L、蔗糖转化率高达50%,并且Levan蔗糖酶合成Levan过程中酶的活性受到pH值、温度、螯合剂、金属离子等多种因素的影响,可以通过控制反应条件促进多糖合成反应的进行。因此,酶法合成将是工业化获得Levan果聚糖的主要方式。     

Rational engineering of natural polyhydroxyalkanoates producing microorganisms for improved synthesis and recovery

Microbial production of biopolymers derived from renewable substrates and waste streams reduces our heavy reliance on petrochemical plastics. One of the most important biodegradable polymers is the family of polyhydroxyalkanoates (PHAs), naturally occurring intracellular polyoxoesters produced for decades by bacterial fermentation of sugars and fatty acids at the industrial scale. Despite the advances, PHA production still suffers from heavy costs associated with carbon substrates and downstream processing to recover the intracellular product, thus restricting market positioning. In recent years, model-aided metabolic engineering and novel synthetic biology approaches have spurred our understanding of carbon flux partitioning through competing pathways and cellular resource allocation during PHA synthesis, enabling the rational design of superior biopolymer producers and programmable cellular lytic systems. This review describes these attempts to rationally engineering the cellular operation of several microbes to elevate PHA production on specific substrates and waste products. We also delve into genome reduction, morphology, and redox cofactor engineering to boost PHA biosynthesis. Besides, we critically evaluate engineered bacterial strains in various fermentation modes in terms of PHA productivity and the period required for product recovery.     

Microbial biosynthesis and secretion of l-malic acid and its applications

l-Malic acid has many uses in food, beverage, pharmaceutical, chemical and medical industries. It can be produced by one-step fermentation, enzymatic transformation of fumaric acid to l-malate and acid hydrolysis of polymalic acid. However, the process for one-step fermentation is preferred as it has many advantages over any other process. The pathways of l-malic acid biosynthesis in microorganisms are partially clear and three metabolic pathways including non-oxidative pathway, oxidative pathway and glyoxylate cycle for the production of l-malic acid from glucose have been identified. Usually, high levels of l-malate are produced under the nitrogen starvation conditions, l-malate, as a calcium salt, is secreted from microbial cells and CaCO₃ can play an important role in calcium malate biosynthesis and regulation. However, it is still unclear how it is secreted into the medium. To enhance l-malate biosynthesis and secretion by microbial cells, it is very important to study the mechanisms of l-malic acid biosynthesis and secretion at enzymatic and molecular levels.