Biochemical Properties and Structure Analysis of a DAG-Like Lipase from Malassezia globosa
Abstract
:1. Introduction
2. Results and Discussion
2.1. Sequence Analysis of MgMDL2
2.2. Expression and Purification of MgMDL2
2.3. Biochemical Properties of MgMDL2
2.3.1. Effect of Temperature on Lipase Activity and Thermostability
2.3.2. Effect of pH on Lipase Activity and Stability
2.3.3. Effect of Chemicals and Organic Solvents on MgMDL2 Activity
Reagents | Relative Activity (%) a | |
---|---|---|
5 mM | 30% (v/v) | |
Control | 100 | 100 |
Ni2+ | 36 ± 1 | - |
Zn2+ | 61 ± 4 | - |
Mn2+ | 35 ± 4 | - |
Mg2+ | 53 ± 4 | - |
Fe2+ | 48 ± 2 | - |
Ca2+ | 42 ± 1 | - |
EDTA | 68 ± 1 | - |
Methanol | - | 98 ± 2 |
Ethanol | - | 97 ± 4 |
Acetone | - | 85 ± 4 |
Isopropanol | - | 41 ± 2 |
2.3.4. Substrate Specificity of MgMDL2
2.4. Molecular Basis for Substrate Selectivity of MgMDL2
3. Experimental Section
3.1. Strains, Plasmids, Chemicals and Materials
3.2. Vector Construction and Transformation of P. pastoris
3.3. Expression and Purification of Recombinant Enzyme
3.4. Biochemical Characterization of Recombinant Enzyme
3.4.1. Enzyme Activity Determination
3.4.2. Determining Temperature-Optimum of Activity and Thermostability of Lipase
3.4.3. Determining pH-Optimum of Activity and pH Stability of Lipase
3.4.4. Substrates Specificity
3.4.5. Effect of Metal Ions on the Enzyme Activity
3.4.6. Effect of Organic Solvents on the Enzyme Activity
3.4.7. Detection of Glycolysation Modification
3.5. Sequence and Structure Analysis
3.5.1. Sequence Analysis
3.5.2. Construction of the Structure in the Open Conformation
3.5.3. Construction of a Model in Complex with Substrate Analogue
3.5.4. MD Simulations
4. Conclusions
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Xu, H.; Lan, D.; Yang, B.; Wang, Y. Biochemical Properties and Structure Analysis of a DAG-Like Lipase from Malassezia globosa. Int. J. Mol. Sci. 2015, 16, 4865-4879. https://doi.org/10.3390/ijms16034865
Xu H, Lan D, Yang B, Wang Y. Biochemical Properties and Structure Analysis of a DAG-Like Lipase from Malassezia globosa. International Journal of Molecular Sciences. 2015; 16(3):4865-4879. https://doi.org/10.3390/ijms16034865
Chicago/Turabian StyleXu, Huan, Dongming Lan, Bo Yang, and Yonghua Wang. 2015. "Biochemical Properties and Structure Analysis of a DAG-Like Lipase from Malassezia globosa" International Journal of Molecular Sciences 16, no. 3: 4865-4879. https://doi.org/10.3390/ijms16034865