Co-evolution of β-glucosidase activity and product tolerance for increasing cellulosic ethanol yield
| العنوان: | Co-evolution of β-glucosidase activity and product tolerance for increasing cellulosic ethanol yield |
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| المؤلفون: | Kexin Wang, Li Hanxin, Xihua Zhao, Huang Qiuxia |
| المصدر: | Biotechnology Letters. 42:2239-2250 |
| بيانات النشر: | Springer Science and Business Media LLC, 2020. |
| سنة النشر: | 2020 |
| مصطلحات موضوعية: | Models, Molecular, 0106 biological sciences, 0301 basic medicine, Bioengineering, Molecular Dynamics Simulation, Protein Engineering, 01 natural sciences, Applied Microbiology and Biotechnology, Evolution, Molecular, Fungal Proteins, 03 medical and health sciences, Hydrolysis, Molecular dynamics, chemistry.chemical_compound, Catalytic Domain, 010608 biotechnology, β glucosidase, Ethanol, beta-Glucosidase, Penicillium, General Medicine, New variant, Nitrophenylgalactosides, Molecular Docking Simulation, Kinetics, Crystallography, 030104 developmental biology, chemistry, Cellulosic ethanol, Yield (chemistry), Product (mathematics), Mutation, Protein Binding, Biotechnology |
| الوصف: | 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 ( $${k}_{cat}/{K}_{m}$$ ) for hydrolyzing pNPG and reduced strength of inhibition ( $${K}_{m}^{app}/{K}_{I}$$ ) by glucose. To be specific, L-6C achieved a $${K}_{m}^{app}/{K}_{I}$$ 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. |
| تدمد: | 1573-6776 0141-5492 |
| DOI: | 10.1007/s10529-020-02935-9 |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::0dec1f859db53d16957cdaf380ea7407 https://doi.org/10.1007/s10529-020-02935-9 |
| Rights: | CLOSED |
| رقم الانضمام: | edsair.doi.dedup.....0dec1f859db53d16957cdaf380ea7407 |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 15736776 01415492 |
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| DOI: | 10.1007/s10529-020-02935-9 |