氨基酸发酵生产的研究进展

由于氨基酸在食品、饲料、医药、农业和日用化工等方面有极其广泛的用途,尤其随着抗癌药物制剂、氨基酸输液制剂及甜味二肽生产的飞速发展,对原料氨基酸的需求量日益增长。传统的发酵工业越来越不能满足需求,势必被以基因工程为基础的新兴发酵工业所代替。通过对发酵法生产氨基酸的历史进行回顾,及对未来前景作出展望,指出了运用ＤＮＡ 重组、定向突变等手段,对代谢途径及关键酶进行了深入系统的研究的必要性。     

氨基酸技术发展及新产品开发

<正>氨基酸及其衍生物具有非常重要的生理功能。氨基酸工业是发酵工业的支柱产业之一,其产品有着广泛的应用和巨大的市场。近些年,氨基酸工业的发展日新月异,各种氨基酸生产的新菌种、新工艺和新技术层出不穷,这为氨基酸工业的进一步发展提供了巨大的动力。主要介绍氨基酸代谢工程的技术发展和氨基酸深层次加工及新产品开发进展。     

氨基酸发酵生产及其产生菌研究进展

<正> 氨基酸是构成蛋白质的基本单位,为生命体所必需。氨基酸的生产主要通过发酵途径来进行。因此高产菌株的选育是氨基酸发酵生产的基础,也是氨基酸工业化生产的重要环节。为保证人类对氨基酸的需求及其在食品、工业、农业和畜牧业上得到广泛应用,就必须加强对氨基酸的生产及其开发利用、以及高效优质生产菌的选育等方面的基础和应用基础的研究,这是氨基酸工业化生产所不可缺少的。     

发酵工业菌种的迭代创制

生物发酵是以微生物菌种为生物催化剂,以淀粉糖、生物质等可再生资源为原料发酵生产各种食品、化学品、燃料、材料等物质的生产过程,具有绿色、低碳和可持续等特征。我国拥有全球规模最大的生物发酵产业,尤其氨基酸、维生素等传统发酵产品占全球市场份额的60%–80%。发展生物发酵产业对于我国实现“碳中和、碳达峰”的战略目标和生物经济发展具有重要的意义。微生物工业菌种是生物发酵产业的核心,直接影响原料路线、产品种类和生产成本。创新发酵工业菌种,提升其原料转化利用效率,提高产物生产水平,拓展产品种类,是生物发酵产业高质量发展的关键。近年来,合成生物学、系统生物学等学科的发展,进一步加深了研究者对微生物底盘细胞生理代谢机制的理解,加速了基因编辑等菌种设计创制使能技术的发展,为发酵工业菌种改造提升提供了新动能。本文选取了具有代表性的大宗氨基酸、B族维生素、柠檬酸、燃料乙醇等发酵产业,从其工业微生物底盘的基础研究和技术开发角度,综述发酵工业菌种改造提升的最新进展,并展望人工智能、自动化与生命科学交叉融合将对工业菌种迭代产生的重要影响。     

利用多菌种混合发酵降解饼粕中粗蛋白的实验研究

对凝结芽孢杆菌、圆孢芽孢杆菌、短小芽孢杆菌和球形芽孢杆菌在菜籽饼粕中混合发酵降低其大分子粗蛋白含量提高游离氨基酸含量的发酵工艺进行了研究。结果表明其最佳发酵工艺条件：4种菌种接种量比例为(1：1：3：1),含水量60％,接种量15％,发酵25d,粗蛋白降解率达41．98％,游离氨基酸含量提高10．18倍。为工业上生产菌肥提供了理论基础。     

氨基酸生产的代谢工程研究进展与发展趋势

氨基酸发酵是我国发酵工业的支柱产业,近年来,随着代谢工程的快速发展,氨基酸的代谢工程育种蓬勃发展。传统的正向代谢工程、基于组学分析与计算机模拟的反向代谢工程以及借鉴自然进化的进化代谢工程,都有越来越多的应用。在氨基酸的工业生产中涌现出了一系列具有高效生产、抗逆性强等优良性状的菌株。日益剧烈的市场竞争对菌株的选育提出了新的要求,如开发高附加值氨基酸品种、菌株代谢的动态调控、适应新工艺的要求等。文中介绍了氨基酸生产相关的代谢工程研究进展以及未来的发展趋势。     

纪念我国发酵法生产味精四十周年--回忆617短杆菌谷氨酸发酵的研究

综观我国当代发酵法生产氨基酸、核苷酸、有机酸、酶制剂、抗生素等研制过程,基本上是跟踪国外先进技术并加以仿制,本文中L-谷氨酸发酵的研制可以说是一个典型范例。从某种意义上来说,如今我国氨基酸和有机酸发酵之发达,莫不始于L-谷氨酸发酵的成功。然而反思我国的发酵工业,虽然L-谷氨酸、柠檬酸、啤酒的产量堪称世界第一,但充其量中国只能称之为发酵工业大国。因此,今后如果不进行自主科技创新,中国的发酵工业就不可能成为真正意义上科技领先的强国。     

工业生物发酵过程模拟：进展与发展趋势

工业生物发酵是工业生物技术规模化生产必需的基本操作单元。对微生物细胞及其反应器进行数学模拟将有助于加深对发酵过程的理解,也将为新的合成生物构建提供解决策略。文中对工业发酵系统的特点、数学模拟的发展历史、数学模型的分类和特点、用途等作了深入阐述,并展望了全发酵系统模拟的发展趋势。     

氨基酸发酵工业的发展近况

一、概述 氨基酸广泛应用于食品、医药、饲料等部门。自1957年日本首创用微生物发酵法工业生产L-谷氨酸以来,各种氨基酸发酵的研究开发蓬勃发展起来,现在大部分氨基酸均可用微生物方法生产了,氨基酸工业已成为新兴的工业部门。据估计1985年世界市场的销售额已超过     

回归创业宗旨谋求发展 降低人工成本的方案也即将出台

以氨基酸发酵．抗体技术等生物科技为核心而发展壮大起来的协和发酵工业株式会社,围绕“以技术革新回报社会”这一创业宗旨进行了企业重组,以谋求今后的发展并进一步深化投资和提高生产效率。[编者按]     

不同抗冷性水稻中编码甘油-3-磷酸转酰酶的部分cDNA的序列比较研究

采用ＲＴ－ＰＣＲ技术,以根据国外报道的几种双子叶植物的甘油－３－磷酸转酰酶的桎保守的氨基酸而设计的简并引物作为扩增引物,从不同抗冷性水稻品种中均扩增到一段约３１５ｂｐ的ｃＤＮＡ片段。测序结果表明它们都是编码ＧＰＡＴ的部分ｃＤＮＡ含有３１５个核苷酸,编码１０５个氨基酸。比产它们之间的核苷酸及推导氨基酸序列,发现有一定差异,且抗冷性相差越大的品种间差异越大。抗冷性的差异可能与脯氨枝的替换有关。     

传染性法氏囊病病毒中国强毒株A节段cDNA基因的克隆和序列分析

以来自哈尔滨传染性法氏囊病病毒（ＩＢＤＶ） 强毒株（Ｈａｒｂｉｎ 毒株,Ｈ） 的基因组ＲＮＡ为模板,用反转录聚合酶链反应（ＲＴ－ ＰＣＲ） 的方法得到了其Ａ 节段的全长ｃＤＮＡ 片段,分５＇端（１ ６５９ｂｐ） 和３＇端（１ ４４４ｂｐ） 上下两段分别克隆到ｐＧＥＭＢ○Ｒ － Ｔ 载体上,测定了其核苷酸顺序,在长为３ １０１ ｂｐ 中含有两个阅读框ＯＲＦＡ１ 和ＯＲＦＡ２ ,分别编码１ ０１２ 个氨基酸的前体蛋白（ＶＰ２ － ４ －３） 和１４５ 个氨基酸的ＶＰ５,ＯＲＦＡ１ 和ＯＲＦＡ２ 有部分的重叠。将核苷酸序列及推测出的氨基酸序列与已报道的ＩＢＤＶ 血清Ⅰ型和Ⅱ型毒株的相应序列进行了比较,结果表明：Ｈ 毒株与其它血清Ⅰ型毒株之间,在核苷酸水平上存在２５ｂｐ － ２６７ｂｐ 的差异;在氨基酸水平上存在１７ ～４０ 个氨基酸的差异。在ＶＰ２ － ４ － ３ 内比较显示,Ｈ 毒株与Ｐ２ 、Ｃｕ－ １ 之间氨基酸的差异最小为１ ．７％ ,Ｈ 毒株与ＵＫ６６１ 之间氨基酸的差异最大为３ ．９ ％ 。变异主要发生在ＶＰ２ 的可变区（２０６ － ３５０ 位氨基酸） ,在Ｈ 毒株所特有的１２ 个氨基酸当中,该区就占５ 个,代表１ ．７６ ％ 的变异。ＶＰ４、ＶＰ３ 和ＶＰ５区各有     

Linkage of Amino Acid Variation and Evolution of Human Immunodeficiency Virus Type 1 gp120 Envelope Glycoprotein (Subtype B) with Usage of the Second Receptor

To clarify the relationship between the amino acid variations of the gp120 of human immunodeficiency virus type 1 (HIV-1) and the chemokine receptors that are used as the second receptor for HIV, we evaluated amino acid site variation of gp120 between the X4 strains (use CXCR4) and the R5 strains (use CCR5) from 21 sequences of subtype B. Our analysis showed that residues 306 and 322 in the V3 loop and residue 440 in the C4 region were associated with usage of the second receptor. The polymorphism at residue 440 is clearly associated with the usage of the second receptor: The amino acid at position 440 was a basic amino acid in the R5 strains, and a nonbasic and smaller amino acid in the X4 strains, while the V3 loop of the X4 strains was more basic than that of the R5 strains. This suggests that residue 440 in the C4 region, which is close to the V3 loop in the three-dimensional structure, is critical in determining which second receptor is used. Analysis of codon frequency suggests that, in almost all cases, the difference at residue 440 between basic amino acids in the R5 strains and nonbasic amino acids in the X4 strains could be due to a single nucleotide change. These findings predict that the evolutionary changes in amino acid residue 440 may be correlated with evolutionary changes in the V3 loop. One possibility is that a change in electric charge at residue 440 compensates for a change in electric charge in the V3 loop. The amino acid polymorphism at position 440 can be useful to predict the cell tropism of a strain of HIV-1 subtype B.     

Molecular analysis of an acatalasemic mouse mutant

The Csb acatalasemia mouse mutant differentially expresses reduced levels of catalase activity in a tissue specific manner. In order to pinpoint the molecular lesion that imparts the acatalasemia phenotype in Csb mice we have utilized the polymerase chain reaction technique to isolate catalase cDNA clones from control and Csb mouse strains. Sequence analyses of these cDNA clones have revealed a single nucleotide difference within the coding region of catalase between control and Csb mice. This nucleotide transversion (G----T) is located in the third position of amino acid 11 in the catalase monomer. In control mouse strains glutamine (CAG) is encoded at amino acid 11, while in Csb mice this codon (CAT) encodes histidine. This amino acid is located within a region that forms the first major alpha-helix in the amino-terminal arm of the catalase subunit and, as such, may render the catalase molecule unstable under certain physiological conditions.     

Evolution of alkaline phosphatase in marine species of Vibrio.

The evolution of alkaline phosphatase was studied in marine species of Vibrio. Two antisera prepared against purified alkaline phosphatases from Vibrio splendidus and Vibrio harveyi were used to estimate the amino acid sequence divergence of this enzyme in 51 strains belonging to nine species. The methods used were the quantitative microcomplement fixation technique and the Ouchterlony double-diffusion procedure. There was a high degree of congruence between the measurement of the amino acid sequence divergence of alkaline phosphatase and the percentage of deoxyribonucleic acid homology of the different organisms relative to both reference strains (correlation coefficient of -0.89) as well as between the amino acid sequence divergence of alkaline phosphatase and superoxide dismutase (correlation coefficient of 0.92) relative to V. splendidus. These findings supported the view that the evolution of marine species of Vibrio is primarily vertical and that horizontal evolution (involving genetic exchange between species), if significant, is restricted to a minor fraction of the bacterial genome.     

Negative interactions between amino acid and methylamine/ammonia transport systems of Saccharomyces cerevisiae.

The transport of methylamine (methylammonium ion) and ammonia (ammonium ion) is accomplished in Saccharomyces cerevisiae by means of a specific active transport system. L-Amino acids are noncompetitive inhibitors of methylamine transport. This inhibition is relieved or eliminated in mutant strains that have a reduced ability to transport amino acids. The inhibition of methylamine transport occurs immediately upon the addition of amino acids to the assay system and persists until the external amino acid pool is depleted. The degree of inhibition observed is a direct function of the rate of amino acid transport. Both methylamine and ammonia are capable of inhibiting amino acid transport. The inhibition of amino acid transport is eliminated in mutant strains that cannot transport methylamine and ammonia.     

Correlations between amino acid concentrations in brains of seizure-susceptible and seizure-resistant rats

We have examined the relationship between free amino acid concentrations in the brains of genetically seizure-susceptible and seizure-resistant rats. The concentrations of free amino acids in the two strains do not differ significantly in the inferior colliculus or the cortex. However, animal-to-animal variations in the concentrations of numerous amino acid pairs are highly correlated. Glutamic acid decarboxylase activities did not vary between the two strains.We conclude that the strong correlations reported between glutamate and taurine levels in several species are not unique to this amino acid pair. Furthermore, unlike the situation with some experimentally-induced epilepsies, genetic epilepsy is not associated with major disturbances in free amino acid concentrations. The high correlations between amino acid pairs in some cases may reflect variations in cellular and subcellular compartment sizes that are shared by several amino acids.     

Oenococcus oeni preference for peptides: qualitative and quantitative analysis of nitrogen assimilation

Optimization of malolactic fermentation in wine depends mainly on better understanding of nitrogen nutritional requirements of Oenococcus oeni. Four widely used starter strains and the reference ATCC BAA-1163 strain were grown in media containing different N sources: free amino acids, oligopeptides (0.5–10 kDa) or polypeptides (> 10 kDa). Amino acid auxotrophies were determined by the single omission technique. The tested strains were indifferent to only two to four amino acids and two of the starter strains appeared to be particularly demanding. Nitrogen consumption was investigated and a significant level of nitrogen was consumed by O. oeni only in the free amino acid medium. In media containing complex nitrogen sources, a global balance above 5 mg N l⁻¹ was enough to ensure biomass formation of all tested strains. Moreover, for all strains, bacterial growth yield was higher in the presence of nitrogen from peptides than that from free amino acids. However, no direct relationship between the bacterial growth level and the amount of nitrogen metabolized could be established. These findings were discussed in relation to the physiology of wine malolactic bacteria.