麦长管蚜唾液中几种酶的鉴定、活力测定与功能分析

用Parafilm膜夹营养液法,以两种食料介质饲喂麦长管蚜Macrosiphum avenae 3龄若蚜并收集其唾液,对唾液中的酶类进行了鉴定、活力测定和功能分析。结果表明,在20%蔗糖介质提取液中,鉴定有果胶酶、多酚氧化酶和纤维素酶; 在水介质提取液中鉴定有纤维素酶; 两种介质提取液中都未鉴定出过氧化物酶。酶活力测定结果表明, 在20%蔗糖介质提取液中, 每30头蚜虫分泌的果胶酶、多酚氧化酶和纤维素酶的酶活力分别为2.59×10^-3 U/g、7×10^-3 U/g和7.89×10^-3 U/g; 在水介质提取液中,纤维素酶活力为3.68×10^-3 U/g。行为反应试验结果表明,果胶酶处理麦苗的挥发物组分能引起麦长管蚜寄生性天敌燕麦蚜茧蜂Aphidius avenae和捕食性天敌七星瓢虫Coccinella septempunctata 的嗅觉偏好反应,因此,果胶酶在麦长管蚜取食诱导小麦植株的间接防御反应中具有重要作用。     

Streptomyces sp. Z-11-6, a novel producer of extracellular L-glutamate oxidase

Glutamate oxidase activity was studied in 1254Streptomyces strains isolated from the zonal soils of various regions of Russia and other countries. Seven strains proved to be producers of extracellular L-glutamate oxidase. The most active producer strain was identified, and the conditions of enzyme biosynthesis were optimized. A multistep mutagenesis-selection procedure allowed a genetically stable strain,Streptomyces sp. Z-11-6, to be obtained, whose glutamate oxidase activity was 40 times higher than that of the original natural isolate.     

Comparative study of hydrogel-immobilized l-glutamate oxidases for a novel thick-film biosensor and its application in food samples

Novel, thick-film biosensors have been developed for the determination of l-glutamate in foodstuffs. The sensors were prepared by immobilization of l-glutamate oxidase by using polycarbamylsulfonate-hydrogel on a thick-film sensor. l-Glutamate oxidases obtained from Streptomyces sp. with different degree of purification were compared with their characteristic response to l-glutamate at different conditions and for their specificity, inhibition, and storage properties. These sensors were applied to determine monosodium glutamate in soy sauce samples and show good correlation with colorimetric method.     

Isolation and purification of methyl mercaptan oxidase fromRhodococcus rhodochrous for mercaptan detection

Methyl mercaptan oxidase was successfully induced fromRhodococcus rhodochrous IGTS8 using methyl mercaptan gas and purified to homogeneity for the detection of mecrcaptans. The purification procedure involved DEAE-Sephacel and Superose 12 column chromatography with recovery yields of 85.8 and 83.3%, and a specific activity of 92.7 and 303.4 units/mg-protein, respectively. The molecular weight of purified methyl mercaptan, oxidase was determined to be 64.5 kDa by SDS-PAGE. The extract from gel filtration chromatography oxidizes methyl mercaptan to produce formaldehyde, which can be easily detected by the purpald-coloring method. Optimum temperature for activity was achieved at 60°C. This enzyme was inhibited by both K₂SO₄ and NaCl at concentration of less than 100 mM and recovered to original activity at concentration of 200 mM. In the presence of methanol, the activity decreased by 33%.     

乳酸氧化酶研究进展

乳酸氧化酶能够氧化乳酸生成丙酮酸,由于反应过程中不需要外源辅酶作为电子受体,而具有较好的应用前景。乳酸氧化酶和许多黄素蛋白酶相比较,具有明显的共性,因此可视为黄素蛋白家族中的一员。乳酸单加氧酶的催化机理和乳酸氧化酶相似,但产物不同,这主要是由于中间产物复合体稳定性的差别。乳酸单加氧酶催化形成的中间复合体EFMNH2pyruvate很稳定,在氧的作用下,生成EFMNpyruvateH2O2中间体,继续反应形成乙酸,CO2和H2O;乳酸氧化酶生成的EFMNH2pyruvate复合体不稳定,丙酮酸很快从复合体上分离下来,还原型中间体EFMNH2被氧氧化,同时形成过氧化氢。     

Biochemical and Histological Changes during In Vitro Organogenesis in Jatropha Integerrima

Biochemical changes associated with adventitious shoot regeneration during in vitro culture of hypocotyl explants of Jatropha integerrima were determined. Histological and biochemical studies were undertaken at 7-d intervals, up to four weeks on hypocotyl explants cultured on basal Murashige and Skoog's medium supplemented with 0.5 mg dm^-3 N⁶-benzyladenine and 1.0 mg dm^-3 indole-3-butyric acid. Initial cell proliferation occurred within one week of culture; meristemoid differentiation within two to three weeks and shoot development after four weeks. Peak activities of alkaline phosphatase, peroxidase and polyphenol oxidase was observed at day 14 indicating their involvement in the formation of meristematic centers. Protein accumulation and acid phosphatase activity were maximum at day 28.     

乙酸合成途径阻断及NADH氧化酶表达对于谷氨酸棒杆菌生产乙偶姻的影响

【目的】研究乙酸合成途径阻断及NADH氧化酶表达对于谷氨酸棒杆菌生产乙偶姻的影响。【方法】在谷氨酸棒杆菌CGF2中异源表达als SD操纵子构建乙偶姻生产菌株CGT1,考察敲除乙酸生成途径cat和pqo对乙偶姻的影响。然后引入短乳杆菌的NADH氧化酶,在优化的溶氧条件下研究其对乙偶姻产量的影响。【结果】CGT1在摇瓶发酵中可积累6.27 g/L乙偶姻,敲除cat使乙偶姻产量显著提高30.94%,达到8.21 g/L;双敲除cat和pqo没有进一步提高产量。通过优化发酵的溶氧水平,乙偶姻产量达到10.06 g/L。在高溶氧水平下引入NADH氧化酶导致菌株的生长和糖代谢速率提高,但乙偶姻产量略有降低。在分批补料发酵中,重组菌株乙偶姻产量达到40.51 g/L,产率为0.51 g/(L?h)。【结论】在谷氨酸棒杆菌中阻断乙酸合成途径cat能够有效提高乙偶姻产量,NADH氧化酶在高溶氧水平下表达不利于乙偶姻的合成,需要进一步调节表达水平以确定其效果。     

彩绒革盖菌固体发酵生产木质素酶工艺优化研究*

采用均匀设计U15(5^8)和双温度培养法进行彩绒革盖菌固体发酵生产木质素酶,对漆酶、愈创木酚酶、多酚氧化酶的活力进行回归分析。结果表明：应用双温度培养法进行彩绒革盖菌固体发酵生产木质素酶时,在自然补给氧气,培养基pH自然(约6．5),并保持环境湿度约60％的条件下,20d是适宜的发酵周期;玉米浆、麸皮、(NH4)2SO4、水分适宜作为固体培养基的成分;少量的Tween-80有利于木质素酶的生产。     

一株海洋低温葡萄糖氧化酶菌株的筛选、鉴定及部分酶学性质

【目的】从海洋样品中分离筛选出产葡萄糖氧化酶菌株。【方法】采用双层平板筛选法进行初筛、复筛确定一株酶活较好的菌株,命名为GOD2(Glucose oxidase)。通过形态学、生理生化特征及16S rRNA基因序列分析研究其分类地位,并对其产生的葡萄糖氧化酶进行分离纯化和部分酶学性质的研究。【结果】细菌GOD2为产葡萄糖氧化酶菌株且遗传稳定,初步鉴定该菌株为假单胞杆菌(Pseudomonas migulae),其所产酶最适反应温度为20°C,热稳定性较差,40°C剩余相对酶活80%;超过40°C酶活力迅速下降。【结论】GOD2是一株极具研究价值的产低温葡萄糖氧化酶菌株。目前没有关于利用该菌生产葡萄糖氧化酶的报道。