Effects of Module Truncation of a New Alginate Lyase VxAly7C from Marine Vibrio xiamenensis QY104 on Biochemical Characteristics and Product Distribution
Abstract
:1. Introduction
2. Results
2.1. Cloning and Sequence Analysis of the Alginate Lyase Gene
2.2. Heterologous Expression and Purification of Recombinant VxAly7C-FL, VxAly7C-TM1, and VxAly7C-TM2
2.3. Biochemical Characteristics of Recombinant VxAly7C-FL, VxAly7C-TM1, and VxAly7C-TM2
2.4. Effects of NaCl, Metal Ions, Chelators, and Detergents on Alginate Lyase Activity
2.5. Substrate Specificity of Recombinant VxAly7C and Its Truncated Mutants
2.6. Analysis of the Mode of Action and End Product Distribution
3. Discussion
4. Materials and Methods
4.1. Bacterial Strains, Plasmids, and Chemicals
4.2. Sequence Analysis
4.3. Construction of Recombinant VxAly7C and Its Truncated Mutants
4.4. Heterologous Expression and Purification of Recombinant Alginate Lyase
4.5. Enzyme Activity and Kinetic Parameter Assay
4.6. Biochemical Characteristics of Recombinant VxAly7C-FL and Its Truncated Mutants
4.7. Substrate Specificity, Mode of Action, and End Product Distribution Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Protein | Specific Activity (U/mg) | Molecular Weight (kDa) | Specific Activity (U/nmol) |
---|---|---|---|
VxAly7C-FL | 557.82 | 72.47 | 40.41 |
VxAly7C-TM1 | 946.34 | 56.72 | 53.73 |
VxAly7C-TM2 | 1351.47 | 40.41 | 54.62 |
VxAly7C-FL | VxAly7C-TM1 | VxAly7C-TM2 | |
---|---|---|---|
Km (mM) | 8.91 ± 0.11 | 12.09 ± 0.07 | 19.23 ± 0.16 |
kcat (s−1) | 1734.86 ± 10.49 | 5523.81 ± 12.31 | 15,960 ± 5.14 |
kcat/Km (s−1·mM−1) | 194.93 ± 2.25 | 456.89 ± 2.45 | 829.95 ± 9.77 |
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Tang, L.; Bao, M.; Wang, Y.; Fu, Z.; Han, F.; Yu, W. Effects of Module Truncation of a New Alginate Lyase VxAly7C from Marine Vibrio xiamenensis QY104 on Biochemical Characteristics and Product Distribution. Int. J. Mol. Sci. 2022, 23, 4795. https://doi.org/10.3390/ijms23094795
Tang L, Bao M, Wang Y, Fu Z, Han F, Yu W. Effects of Module Truncation of a New Alginate Lyase VxAly7C from Marine Vibrio xiamenensis QY104 on Biochemical Characteristics and Product Distribution. International Journal of Molecular Sciences. 2022; 23(9):4795. https://doi.org/10.3390/ijms23094795
Chicago/Turabian StyleTang, Luyao, Mengmeng Bao, Ying Wang, Zheng Fu, Feng Han, and Wengong Yu. 2022. "Effects of Module Truncation of a New Alginate Lyase VxAly7C from Marine Vibrio xiamenensis QY104 on Biochemical Characteristics and Product Distribution" International Journal of Molecular Sciences 23, no. 9: 4795. https://doi.org/10.3390/ijms23094795