Characterization and Catalytic-Site-Analysis of an Aldo-Keto Reductase with Excellent Solvent Tolerance
Abstract
:1. Introduction
2. Results
2.1. Cloning and Sequencing Analysis of AKR3-2-9
2.2. Expression and the Purification of AKR3-2-9
2.3. Substrate Specificity
2.4. Effect of Temperature and pH
2.5. Detection of AKR3-2-9 Tolerant Organic Solvent Properties
2.6. Kinetic Analysis
2.7. The Activity towards DKTP
2.8. Molecular Modeling
3. Discussion
4. Materials and Methods
4.1. Strains, Vectors, Chemicals
4.2. Homologous Protein-Searching Analysis and Homology Modeling
4.3. Cloning, Expression and Purification of AKR3-2-9
4.4. Enzyme Activity Assays and Kinetic Parameter Studies
4.5. Characterization of the AKR 3-2-9
4.5.1. Substrate Specificity of AKR3-2-9
4.5.2. Effect of Temperature and pH
4.5.3. Organic Solvent Tolerance of AKR3-2-9
4.6. The Activity towards DKTP
4.7. The GenBank Accession Number of the AKR3-2-9
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Substrate | Km (mM) | kcat (s−1) | kcat/Km (s−1 mM−1) |
---|---|---|---|
Acetylacetone | 6.29 ± 0.36 | (4.195 ± 0.08) × 103 | 0.667 × 103 |
Methyl pyruvate | 7.431 ± 0.47 | (4.24 ± 0.09) × 103 | 0.571 × 103 |
Phenoxyacetone | 9.844 ± 0.82 | (3.648 ± 0.12) × 103 | 0.371 × 103 |
DKTP | 2.81 ± 0.78 | (1.601 ± 0.19) × 103 | 0.569 × 103 |
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Pei, R.; Wu, W.; Zhang, Y.; Tian, L.; Jiang, W.; Zhou, S.-F. Characterization and Catalytic-Site-Analysis of an Aldo-Keto Reductase with Excellent Solvent Tolerance. Catalysts 2020, 10, 1121. https://doi.org/10.3390/catal10101121
Pei R, Wu W, Zhang Y, Tian L, Jiang W, Zhou S-F. Characterization and Catalytic-Site-Analysis of an Aldo-Keto Reductase with Excellent Solvent Tolerance. Catalysts. 2020; 10(10):1121. https://doi.org/10.3390/catal10101121
Chicago/Turabian StylePei, Rui, Weiliang Wu, Yuqian Zhang, Libing Tian, Wei Jiang, and Shu-Feng Zhou. 2020. "Characterization and Catalytic-Site-Analysis of an Aldo-Keto Reductase with Excellent Solvent Tolerance" Catalysts 10, no. 10: 1121. https://doi.org/10.3390/catal10101121