Extracellular Production of a Novel Endo-β-Agarase AgaA from Pseudomonas vesicularis MA103 that Cleaves Agarose into Neoagarotetraose and Neoagarohexaose
Abstract
:1. Introduction
2. Results and Discussion
2.1. Domain Prediction of AgaA from P. vesicularis MA103
2.2. Cloning
2.3. Effect of Temperature on AgaA Overexpression
2.4. Effect of the N-Terminal Signal Peptide on the Overexpression of Functional AgaA
Protein Source * | Total Protein (mg) | Total Activity (units) | Specific Activity (units/mg) |
---|---|---|---|
AgaA-FL | 12.2 | 5192.3 | 425.6 |
AgaA-∆SP | 18.3 | 689.9 | 37.7 |
AgaA-PelB | 6.2 | 816.8 | 132.6 |
2.5. AgaA from P. vesicularis Is an Endo-Type β-Agarase Yielding Neoagarotetraose and Neoagarohexaose Products
3. Experimental Section
3.1. Isolation of Pseudomonas vesicularis MA103
3.2. Cloning of AgaA
3.3. AgaA Overexpression and Purification
3.4. Western Blot
3.5. Enzyme Activity Assay
3.6. Time Course Hydrolysis of Agarose
3.7. Thin-Layer Chromatography (TLC)
3.8. Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS)
3.9. Statistical Analysis
4. Conclusions
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Hsu, P.-H.; Wei, C.-H.; Lu, W.-J.; Shen, F.; Pan, C.-L.; Lin, H.-T.V. Extracellular Production of a Novel Endo-β-Agarase AgaA from Pseudomonas vesicularis MA103 that Cleaves Agarose into Neoagarotetraose and Neoagarohexaose. Int. J. Mol. Sci. 2015, 16, 5590-5603. https://doi.org/10.3390/ijms16035590
Hsu P-H, Wei C-H, Lu W-J, Shen F, Pan C-L, Lin H-TV. Extracellular Production of a Novel Endo-β-Agarase AgaA from Pseudomonas vesicularis MA103 that Cleaves Agarose into Neoagarotetraose and Neoagarohexaose. International Journal of Molecular Sciences. 2015; 16(3):5590-5603. https://doi.org/10.3390/ijms16035590
Chicago/Turabian StyleHsu, Pang-Hung, Chien-Han Wei, Wen-Jung Lu, Fen Shen, Chorng-Liang Pan, and Hong-Ting Victor Lin. 2015. "Extracellular Production of a Novel Endo-β-Agarase AgaA from Pseudomonas vesicularis MA103 that Cleaves Agarose into Neoagarotetraose and Neoagarohexaose" International Journal of Molecular Sciences 16, no. 3: 5590-5603. https://doi.org/10.3390/ijms16035590