通过半理性设计策略和计算机分析提高葡萄糖胺-6-磷酸合酶的酶活性。
Improving enzyme activity of glucosamine-6-phosphate synthase by semi-rational design strategy and computer analysis.
机构信息
State Key Laboratory of Biobased Material and Green Papermaking (LBMP) (Qilu University of Technology), Jinan, 250353, Shandong, People's Republic of China.
Key Laboratory of Shandong Microbial Engineering, QILU University of Technology (Shandong Academy of Sciences), Jinan, 250353, Shandong, People's Republic of China.
出版信息
Biotechnol Lett. 2020 Nov;42(11):2319-2332. doi: 10.1007/s10529-020-02949-3. Epub 2020 Jun 29.
OBJECTIVE
To improve enzyme activity of Glucosamine-6-phosphate synthase (Glms) of Bacillus subtilis by site saturation mutagenesis at Leu, Ala, Lys, Ser and Val based on computer-aided semi-rational design.
RESULTS
The results indicated that LS had the greatest effect on the activity of BsGlms and the enzyme activity increased from 5 to 48 U/mL. The mutation of LS increased the yield of glucosamine by 1.6 times that of the original strain. The binding energy of the mutant with substrate was reduced from - 743.864 to - 768.246 kcal/mol. Molecular dynamics simulation results showed that Ser enhanced the flexibility of the protein, which ultimately led to increased enzyme activity.
CONCLUSION
We successfully improved BsGlms activity through computer simulation and site saturation mutagenesis. This combination of methodologies may fit into an efficient workflow for improving Glms and other proteins activity.
目的
通过基于计算机辅助半理性设计的亮氨酸、丙氨酸、赖氨酸、丝氨酸和缬氨酸的定点饱和突变，提高枯草芽孢杆菌葡萄糖-6-磷酸合酶（Glms）的酶活性。
结果
结果表明，LS 对 BsGlms 的活性影响最大，酶活性从 5 提高到 48 U/mL。LS 的突变使葡萄糖胺的产量增加了 1.6 倍。突变体与底物的结合能从 -743.864 降低到 -768.246 kcal/mol。分子动力学模拟结果表明，丝氨酸增强了蛋白质的灵活性，最终导致酶活性增加。
结论
我们通过计算机模拟和定点饱和突变成功提高了 BsGlms 的活性。这种组合方法可能适合于提高 Glms 和其他蛋白质活性的有效工作流程。
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