Characterization of a Novel Superoxide Dismutase from a Deep-sea Sea Cucumber (Psychoropotes verruciaudatus)
Abstract
:1. Introduction
2. Materials and Methods
2.1. Materials, Total RNA Extraction, and cDNA Cloning
2.2. Sequence Analysis of PVCuZnSOD
2.3. Recombinant Protein Expression
2.4. Extraction and Purification of Recombinant Protein
2.5. Western Blot Analysis
2.6. PVCuZnSOD Assay
2.7. Effect of Temperature on the Activity of PVCuZnSOD
2.8. Effect of pH on the Activity of PVCuZnSOD
2.9. Effect of Metal Ions on the Activity of PVCuZnSOD
2.10. Effect of Surfactants on the Activity of PVCuZnSOD
2.11. Effects of Organic Solvents on the Activity of PVCuZnSOD
2.12. Effect of Denaturant on the Activity of PVCuZnSOD
2.13. Effect of Digestive Enzyme on the Activity of PVCuZnSOD
2.14. Determination of Kinetic Parameters
2.15. Statistical Analysis
3. Results
3.1. Sequence and Homology Analysis of PVCuZnSOD
3.2. Protein Expression and Purification
3.3. Effect of Temperature
3.4. Effect of pH
3.5. Effect of Metal Ions
3.6. Effect of Surfactants
3.7. Effect of Organic Solvents
3.8. Effect of Denaturants
3.9. Effect of Digestive Enzyme
3.10. Kinetic Parameters
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Genbank | Species | Abbreviation |
---|---|---|
OQ687337 | P. verruciaudatus | PVCuZnSOD |
MK580169.1 | Psychropotes longicauda | PLCuZnSOD |
MK580168.1 | Benthodytes marianensis | BMCuZnSOD |
MG989672.1 | Paelopatides sp. | PACuZnSOD |
JX097096.1 | Apostichopus japonicus | AJCuZnSOD |
KT594770.1 | Sterechinus neumayeri | SNCuZnSOD |
MF324875.1 | Eisenia fetida | EFCuZnSOD |
KR106132.1 | Eisenia andrei | EACuZnSOD |
FM177867.1 | Mytilus galloprovincialis | MGCuZnSOD |
AY332385.2 | Lymnaea stagnalis | LSCuZnSOD |
JX470524.1 | Pelodiscus sinensis | PSCuZnSOD |
BT082811.1 | Anoplopoma fimbria | AFCuZnSOD |
JN032591.1 | Amphiprion clarkii | ACCuZnSOD |
AF329278.1 | Pagrus major | PMCuZnSOD |
MH716024.1 | Mugil incilis | MICuZnSOD |
AY491056.1 | Oreochromis mossambicus | OMCuZnSOD |
HQ318825.1 | Lates calcarifer | LCCuZnSOD |
AY613390.1 | Oplegnathus fasciatus | OFCuZnSOD |
MG208871.1 | Nibea albiflora | NACuZnSOD |
MK860772.1 | Planiliza haematocheilus | PHCuZnSOD |
MT123896.1 | Bostrychus sinensis | BSCuZnSOD |
X95726.1 | Brassica juncea | BJCuZnSOD |
DQ481231.1 | Populus suaveolens | POCuZnSOD |
KU240390.1 | Solanum melongena | SMCuZnSOD |
JQ692111.2 | Vitis vinifera | VVCuZnSOD |
Name of Enzyme | No. of Cleavages | Positions of Cleavage Sites |
---|---|---|
Pepsin (pH > 2) | 17 | 8 9 19 20 35 42 47 61 81 82 103 104 114 123 124 132 133 |
Chymotrypsin | 2 | 20 48 |
Trypsin | 10 | 21 34 66 77 94 113 126 141 150 151 |
Residual Activity (%) | |||
---|---|---|---|
Time (h) | Bovine SOD | PVCuZnSOD | |
Pepsin | 0 | 100.00 ± 2.05 | 100.00 ± 0.75 |
1 | 88.21 ± 3.86 | 93.21 ± 0.70 | |
2 | 42.29 ± 4.86 | 81.28 ± 1.43 | |
3 | 2.18 ± 8.29 | 69.59 ± 1.16 | |
Trypsin/chymotrypsin | 0 | 100.00 ± 4.94 | 100.00 ± 2.27 |
1 | 75.79 ± 1.10 | 121.57 ± 2.30 | |
2 | 43.92 ± 4.11 | 105.73 ± 3.29 | |
3 | 17.85 ± 6.20 | 87.96 ± 7.50 |
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Li, Y.; Chen, Z.; Zhang, P.; Gao, F.; Wang, J.; Lin, L.; Zhang, H. Characterization of a Novel Superoxide Dismutase from a Deep-sea Sea Cucumber (Psychoropotes verruciaudatus). Antioxidants 2023, 12, 1227. https://doi.org/10.3390/antiox12061227
Li Y, Chen Z, Zhang P, Gao F, Wang J, Lin L, Zhang H. Characterization of a Novel Superoxide Dismutase from a Deep-sea Sea Cucumber (Psychoropotes verruciaudatus). Antioxidants. 2023; 12(6):1227. https://doi.org/10.3390/antiox12061227
Chicago/Turabian StyleLi, Yanan, Zongfu Chen, Peng Zhang, Feng Gao, Junfeng Wang, Li Lin, and Haibin Zhang. 2023. "Characterization of a Novel Superoxide Dismutase from a Deep-sea Sea Cucumber (Psychoropotes verruciaudatus)" Antioxidants 12, no. 6: 1227. https://doi.org/10.3390/antiox12061227
APA StyleLi, Y., Chen, Z., Zhang, P., Gao, F., Wang, J., Lin, L., & Zhang, H. (2023). Characterization of a Novel Superoxide Dismutase from a Deep-sea Sea Cucumber (Psychoropotes verruciaudatus). Antioxidants, 12(6), 1227. https://doi.org/10.3390/antiox12061227