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131.
Xiaoyan Guo Yunfang Gao Fangzheng Liu Yong Tao Haibo Jin Jianjun Wang Sheng Wu 《Microbial biotechnology》2023,16(6):1333-1343
R-1,3-butanediol (R-1,3-BDO) is an important chiral intermediate of penem and carbapenem synthesis. Among the different synthesis methods to obtain pure enantiomer R-1,3-BDO, oxidation–reduction cascades catalysed by enzymes are promising strategies for its production. Dehydrogenases have been used for the reduction step, but the enantio-selectivity is not high enough for further organic synthesis efforts. Here, a short-chain carbonyl reductase (LnRCR) was evaluated for the reduction step and developed via protein engineering. After docking result analysis with the substrate 4-hydroxy-2-butanone (4H2B), residues were selected for virtual mutagenesis, their substrate-binding energies were compared, and four sites were selected for saturation mutagenesis. High-throughput screening helped identify a Ser154Lys mutant which increased the catalytic efficiency by 115% compared to the parent enzyme. Computer-aided simulations indicated that after single residue replacement, movements in two flexible areas (VTDPAF and SVGFANK) facilitated the volumetric compression of the 4H2B-binding pocket. The number of hydrogen bonds between the stabilized 4H2B-binding pocket of the mutant enzyme and substrate was higher (from four to six) than the wild-type enzyme, while the substrate-binding energy was decreased (from −17.0 kJ/mol to −29.1 kJ/mol). Consequently, the catalytic efficiency increased by approximately 115% and enantio-selectivity increased from 95% to 99%. Our findings indicate that compact and stable substrate-binding pockets are critical for enzyme catalysis. Lastly, the utilization of a microbe expressing the Ser154Lys mutant enzyme was proven to be a robust process to conduct the oxidation–reduction cascade at larger scales. 相似文献
132.
为探究燕麦(Avena sativa)-绿豆(Phaseolus radiatus)间作效应及氮素转移特性, 在不施氮肥的大田试验条件下, 设置3种种植模式(燕麦单作、绿豆单作和燕麦-绿豆间作), 采用传统挖根法和15N同位素标记法进行研究。结果表明, 间作系统中燕麦侵袭力强于绿豆, 绿豆生长受到抑制。整个生育期, 间作燕麦地上部干物质积累量比单作增加14.9%-33.1%, 2年成熟期间作燕麦的氮素积累量比单作分别提高53.1%和44.8%; 间作减少了开花结荚期绿豆氮素积累量和根瘤重量, 降低了绿豆的固氮效率, 绿豆的固氮效率2年平均降低23.7%, 生物固氮量平均减少11.66%。间作绿豆向燕麦的氮素转移率2年平均值达31.7%, 氮素转移量为212.16 kg∙hm-2。燕麦-绿豆间作降低了开花结荚期绿豆的根瘤固氮酶活性和固氮效率, 但绿豆体内氮素转移增加了燕麦对氮素的吸收利用, 实现了地上部与地下部生长的相互调节和促进, 优化了农田生态系统的氮素管理。 相似文献
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136.
M. Guo S. L. Meyer H. Kaur J. J. Gao K. E. Neet 《Protein science : a publication of the Protein Society》1996,5(3):447-455
An understanding of the structure-function relationship of nerve growth factor (NGF) requires precise knowledge of all the residues and regions that participate in NGF receptor binding, receptor activation, and biological activity. Seven recombinant human NGF mutants having alanine substituted for residues located either in the NGF dimer interface or beta-strand region were studied to determine the role of each amino acid residue in NGF biological activity. F86A, T91A, R100A, and R103A remained nearly full active with 61, 120, 91, and 73% of wild-type activity, respectively, in the PC12 cell bioassay. Hydrophobic core and dimer interface residues Y52, F53, and F54 were studied in more detail. Y52A and F54A were expressed in very low levels, suggesting that these two residues may be important for protein stability. Y52A retained full biological activity (91%). F53A had a 20- and 70-fold reduction in biological activity and TrkA phosphorylation, respectively, with only a 5- to 10-fold effect on TrkA binding and no effect on low-affinity receptor binding. F54A had significantly decreased TrkA phosphorylation and biological activity (40-fold). The results suggest that F53 and F54 may play a structural role in TrkA receptor activation subsequent to binding. 相似文献
137.
冬小麦根表面氧化还原活力的研究 总被引:4,自引:1,他引:3
证实了两个不同品种的冬小麦根系表面存在着氧化NADH和还原K3Fe(CN)6的氧化的活力。还原铁氰化物活力在PH5.5到8.5范围内随着PH值升高而增大,温度在15℃到45℃范围内随温度升高还原活力增强,45℃达最高值,55℃时活力急剧下降。 相似文献
138.
研究了层理鞭枝藻藻胆体在不同浓度磷酸缓冲溶液中解离过程中荧光发射光谱的变化和光能传递。完整藻胆体的77K荧光光谱中只有一个峰,位于685nm它是末端发射体(核心-膜连接多肽和别藻蓝蛋白-B)的荧光峰。部分解离藻胆体的荧光光谱的主峰位移至652nm:次峰位于685nm;660nm为一弱荧光发射肩。它们依次为C-藻蓝蛋白,末端发射体和别藻蓝蛋白的荧光。严重解离藻胆体的荧光主峰移644nm;次峰由685nm移至682nm;660nm荧光发射肩消失。这表明C-藻蓝蛋白所捕获的光能已不能传递给别藻蓝蛋白,但可传递给末端发射体洞时又表明C-藻蓝蛋白不仅与别藻蓝蛋白相连接而且还与末端发射体相连接。提出该藻胆体光能传递链如下:核心-膜连接多肽藻红蓝蛋白→C-藻蓝蛋白→别藻蓝蛋白别藻蓝蛋白-B 相似文献
139.
利用32P同位素示踪法对白浆土中P肥利用率进行了研究.结果表明,在岗地白浆土上,表层土壤表现出一定程度的供P不足现象,白浆层土壤有效P含量严重缺乏.表层土壤中当季P肥利用率的变幅范围为6.09~12.35%,白浆层土壤P肥利用率为13%左右.施用有机肥能明显提高白浆土Olsen P的含量,加速土壤本身P的活化,各种有机物中,以猪粪对土壤潜在P的活化效果最好.将Olsen P与X值、A值比较,认为Olsen P是评价白浆上P肥力简便易行的可靠指标. 相似文献
140.
南美藜种皮凝集素的初步研究 总被引:1,自引:0,他引:1
南美藜种皮中存在着凝集素。最适pH7—9,热稳定性较差。南美藜种皮凝集素(QSL)的血凝活性可以被D-半乳糖,L-鼠李糖抑制,其中前者抑制能力强。QSL硫酸铵分级沉淀血凝活性60—80组分最高。 相似文献