黄原胶降解菌的筛选及其降解酶性质的研究*

从野外采集并筛选到一株能降解黄原胶的菌株,并对菌种进行了对照鉴定。此外,对菌株的发酵参数、黄原胶降解酶的性质(最适反应温度、pH值,金属离子的影响、底物专一性、动力学研究等)作了初步的研究。结果显示,该酶的最适催化反应温度和pH分别为30℃～40℃和5.0～7.0;能够专一性降解黄原胶;测试大多数金属离子对酶活力没有明显影响,但Ca²⁺能很大程度地缓解EDTA对酶活的抑制作用。     

新分离Microbacterium sp．XT11菌产黄原胶降解酶生产条件的优化研究

新分离Microbacterium sp．XT11菌能够合成黄原胶降解酶,将植物病原菌野油菜黄单孢菌分泌的毒素因子黄原胶分解,生成具有激发子和抗微生物活性的黄原胶寡糖。实验确认,黄原胶和酵母浸粉分别是XT11菌生产黄原胶降解酶的最适碳源和氮源,获得最高酶活力的最低碳源和氮源浓度均为0.3％。XT11菌生产黄原胶降解酶的最适条件为：培养温度28℃,培养基起始pH7.0,转速150r／min。     

新分离Microbacterium sp. XT11菌产黄原胶降解酶生产条件的优化研究

新分离Microbacteriumsp.XT11菌能够合成黄原胶降解酶,将植物病原菌野油菜黄单孢菌分泌的毒素因子黄原胶分解,生成具有激发子和抗微生物活性的黄原胶寡糖。实验确认,黄原胶和酵母浸粉分别是XT11菌生产黄原胶降解酶的最适碳源和氮源,获得最高酶活力的最低碳源和氮源浓度均为0.3%。XT11菌生产黄原胶降解酶的最适条件为:培养温度28℃,培养基起始pH7.0,转速150r/min。     

黄原胶降解菌BIT-BJ001培养条件的优化

黄原胶在采油工程中有重要的应用,但其难降解性质给采油工程带来很多问题。从塔里木油田胡杨木根部样品中分离得到1株黄原胶降解菌BIT-BJ001,对其发酵条件的研究表明,此黄原胶降解菌最适培养条件为:黄原胶0.3%,酵母粉0.5%,Na+浓度0.8%,Mg2+浓度0.8%,初始pH值为10,温度60℃。菌种BIT-BJ001降解黄原胶的能力与发酵时间、发酵液中还原糖浓度有关,发酵96 h,黄原胶降粘率达到最高,发酵液中还原糖浓度过高,将抑制菌株对黄原胶的降解。     

黄原胶降解及其抗氧化性能研究

酸性条件下对黄原胶进行氧化降解,透析得到两种黄原胶寡糖,对产物进行FT-IR表征,GPC法测定两种寡糖的分子量分别为7500、10100。考察两种产物对超氧阴离子自由基O2-.和过氧化氢的清除活性以及还原能力,结果表明10100-XG较7500-XG具有更强的抗氧化性能。这可能与黄原胶寡糖活性羟基数目有关。     

黄原胶降解的研究进展

综述了黄原胶的理化特性、降解意义及降解方法,重点介绍了生物法降解黄原胶的国内外研究进展,并对降解黄原胶的研究方向提出了寻求更多降解途径、开发寡糖产品等建议.     

黄原胶寡糖生物活性的研究

利用黄原胶降解菌Cellulom onassp.XT11生产的黄原胶降解酶,对黄原胶进行生物降解,生产具有不同粘度/还原末端比的黄原胶寡糖,并研究了黄原胶寡糖在清除羟基自由基、植物防卫反应中激活因子活性和对植物病原菌抑制能力等方面的生物活性,结果表明黄原胶寡糖具有清除羟基自由基能力,并能激活植物防卫系统以抵御病原菌的侵染,同时对野油菜黄单孢菌也具有抑菌活性。     

黄原胶发酵液纯化精制研究

溶剂法直接提取的黄原胶产品含有大量菌体蛋白和色素杂质,总氮含量高、透明度差、色泽暗深。通过中性蛋白酶处理发酵液,使成品含氮量下降５６．５％;通过Ｎａ２ＳＯ３漂白液,使产品色泽大为改善。实验得出最佳酶解条件：发酵液稀释１倍,温度４４℃、ｐＨ７,加酶量１００ｕ／ｇ发酵液,作用时间２．５ｈ;最佳Ｎａ２ＳＯ３漂白条件：温度２０￣３０℃,ｐＨ５￣６,Ｎａ２ＳＯ３用量为１％（ｗ／ｗ）,作用时间１￣１．５ｈ。     

一株耐高温细菌CHB1的分离和产酶特性研究

从土壤中分离到1株耐高温细菌CHB1,经鉴定为嗜热脂肪土芽胞杆菌(Geobacillus stearothermophilus)。通过平板透明圈法研究其产酶特性,结果表明CHB1具有蛋白酶、淀粉酶和纤维素酶活性,产酶最适温度均为60℃;最佳产酶时间因酶的种类而不同,淀粉酶为24 h,蛋白酶和纤维素酶为48 h。培养基厚度对产酶有一定的影响,以每皿20~25 mL为宜。     

氰戊菊酯降解菌FDB的分离鉴定及其生长特性

从长期受农药污染的农田土壤中分离筛选到一株降解氰戊菊酯杀虫剂的细菌菌株FDB。经形态和生理生化特征鉴定以及对16SrDNA序列进行同源性比较,将该菌株鉴定为铜绿假单胞菌Pseudomonas aeruginosa。FDB能以氰戊菊酯杀虫剂为唯一碳源生长,在30°C培养5d对100mg/L氰戊菊酯异构体的降解率分别达到69.06%(SR+RS)和64.32%(SS+RR)。FDB的最适生长条件为:温度35°C,初始pH值7.0,250mL摇瓶装液量75mL。采用超声波方法破碎菌体细胞,得到粗酶液。胞内和胞外粗酶液对氰戊菊酯异构体的降解试验表明,FDB的氰戊菊酯降解酶属于胞内蛋白组分。     

甲基营养菌WB-1甲胺磷降解酶的产生、部分纯化及性质

甲基营养菌WB１菌株在以甲胺磷作碳氮源的无机盐培养基上生长时,产生甲胺磷降解酶情况较好,培养20 h为细胞收获的适宜时期。所得细胞经过超声波破碎、吐温20抽提、热处理(60℃,9min),DEAE|纤维素32和CM|纤维素32柱层析等步骤,得到的酶制品比活力为180U/mg,收率78.8%,纯化倍数22.8。纯化的酶制品经连续聚丙烯酰胺凝胶电泳,呈现一条主带,酶活力染色则呈现一条与之对应的活性谱带;该酶催化反应的适宜pH为90,底物专一性不强,Hg²⁺,Mn²⁺,Cu²⁺对酶有强烈的抑制作用。部分纯化酶制品在-20℃(0.1mol/L的磷酸盐溶液中)存放5d,酶活力丧失约60%。     

Characteristics of the Kabicidin Resistant Mutant of Fusarium sp. Having a High Productivity of Alkaline Protease

In order to elucidate the biochemical mechanism of the alkaline protease accumulation from n-paraffins by a kabicidin-resistant mutant of Fusarium sp., the cell constituents and the extracellular products of the mutant strain were compared with those of the parent strain. No prominent differences in the cell constituents were observed between the parent and the mutant. From the analysis of the extracellular products, however the mutant was found to have a high productivity of some hydrolytic enzymes, such as amylase and ribonuclease, and ergosterol which is a structural constituent of fungal cell membrane. The relationship of secretion of ergosterol, resistance to kabicidin and accumulation of alkaline protease is discussed.     

Purification and properties of a xanthan depolymerase from a heat-stable salt-tolerant bacterial consortium

Summary A bacterial consortium (NRRL B-14401) resulting from soil enrichment growth on xanthan gum produces enzymes that can degrade xanthan gum in salt-containing solutions at temperatures up to 65°C. One component that cleaves the backbone linkages of both xanthan gum and carboxymethyl cellulose is called xanthan depolymerase. Two such depolymerase activities were isolated by high performance anion exchange chromatography, and their molecular weights determined by size exclusion chromatography to be 170 000 and 100 000 Da. The 170-kDa protein was purified and its properties studied. Sodium chloride and potassium chloride enhanced the hydrolysis of carboxymethyl cellulose, but decreased the rate of degradation of xanthan gum. The purified enzyme, which was optimally active at pH 6, was less stable to extremes of temperature than crude mixtures of cell-free culture broth; stabilized by i substrate it was active for more than 6 h at 50°C.The mention of firm names or trade products does not imply that they are endorsed or recommended by the US Department of Agriculture over other firms or similar products not mentioned.     

海洋微生物ZD02信号降解酶基因aiiA克隆及其生物信息学分析

根据细菌群体感应信号降解酶(AiiA)基因设计简并引物,从海洋微生物ZD02基因组中扩增编码AiiA的基因aiiA,筛选其阳性克隆,测序后分析其基因序列.结果表明:筛选到的阳性克隆ZD02-aiiA序列全长753 bp(Genbank登录号:KC756214),存在一个开放阅读框(ORF),编码由250个氨基酸残基组成的多肽,预测分子量为28.1 kDa,蛋白等电点为4.78.由该序列推导得到的氨基酸残基序列含有一个保守结构域(Lactamase-B) (34AA～235AA),并预测了AiiA的三级结构.以上研究为该序列的重组表达及其相关活性研究奠定了基础.     

Enzymes from psychrophilic organisms

Abstract: Psychrophilic organisms such as micro-organisms and other ectothermic species living in polar, deep- sea or any constantly low temperature environments, produce enzymes adapted to function at low temperature. These enzymes are characterized by a high catalytic efficiency at low and moderate temperatures but are rather thermolabile. Due to their high specific activity and their rapid inactivation at temperatures as low as 30°C, they offer, along with the producing micro-organisms, a great potential in biotechnology. The molecular basis of the adaptation of cold α-amylase, subtilisin, triose phosphate isomerase from Antarctic bacteria and of trypsin from fish living in North Atlantic and in Antarctic sea waters have been studied. The comparison of the 3D structures obtained either by protein modelling or by X-ray crystallography (North Atlantic trypsin) with those of their mesophilic counterparts indicates that the molecular changes tend to increase the flexibility of the structure by a weakening of the intramolecular interactions and by an increase of the interactions with the solvent. For each enzyme, the most appropriate strategy enabling it to accommodate the substrate at a low energy cost is selected. There is a price to pay in terms of thermosensibility because the selective pressure is essentially oriented towards the harmonization of the specific activity with ambient thermal conditions. However, as demonstrated by site-directed mutagenesis experiments carried out on the Antarctic subtilisin, the possibility remains to stabilize the structure of these enzymes without affecting their high catalytic efficiency.     

转解毒酶基因工程菌的构建及酶活性分析

实验构建了能高效降解有机磷、有机氯等四大类农药的转解毒酶基因工程菌。将抗性库蚊解毒酶酯酶B1基因片段引入融合表达载体pThioHisA中,转化入大肠杆菌DH5α,在IPTG诱导下,经过8 h,酯酶B1在大肠杆菌中的获得融合高效表达。研究了工程菌的酶酯活性,重组质粒pThioHisA-B1表达的酯酶融合蛋白具有较高的酯酶B1活性,能高效降解酯酶的特异性底物α-乙酸萘酯(-αNA)和β-乙酸萘酯(-βNA),该工程菌将为农药污染的生物治理提供新手段。