| Autor: |
Wang K; College of Life Science, Jiangxi Normal University, Nanchang, 330022, China., Huang Q; College of Life Science, Jiangxi Normal University, Nanchang, 330022, China., Li H; College of Life Science, Jiangxi Normal University, Nanchang, 330022, China., Zhao X; College of Life Science, Jiangxi Normal University, Nanchang, 330022, China. xhzhao@jxnu.edu.cn. |
| Jazyk: |
angličtina |
| Zdroj: |
Biotechnology letters [Biotechnol Lett] 2020 Nov; Vol. 42 (11), pp. 2239-2250. Date of Electronic Publication: 2020 Jun 24. |
| DOI: |
10.1007/s10529-020-02935-9 |
| Abstrakt: |
β-Glucosidase (BGL) plays a key role in cellulose hydrolysis. However, it is still a great challenge to enhance product tolerance and enzyme activity of BGL simultaneously. Here, we utilized one round error-prone PCR to engineer the Penicillium oxalicum 16 BGL (16BGL) for improving the cellulosic ethanol yield. We identified a new variant (L-6C), a triple mutant (M280T/V484L/D589E), with enhanced catalytic efficiency ([Formula: see text]) for hydrolyzing pNPG and reduced strength of inhibition ([Formula: see text]) by glucose. To be specific, L-6C achieved a [Formula: see text] of 0.35 at a glucose concentration of 20 mM, which was 3.63 times lower than that attained by 16BGL. The catalytic efficiency for L-6C to hydrolyze pNPG was determined to be 983.68 mM -1 s -1 , which was 22% higher than that for 16BGL. However, experiments showed that L-6C had reduced binding affinity (2.88 mM) to pNGP compared with 16BGL (1.69 mM). L-6C produced 6.15 g/L ethanol whose yield increased by about 10% than 16BGL. We performed molecular docking and molecular dynamics (MD) simulation, and binding free energy calculation using the Molecular Mechanics/Poisson Boltzmann surface area (MM/PBSA) method. MD simulation together with the MM/PBSA calculation suggested that L-6C had reduced binding free energy to pNPG, which was consistent with the experimental data. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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