Venomics of Trimeresurus (Popeia) nebularis, the Cameron Highlands Pit Viper from Malaysia: Insights into Venom Proteome, Toxicity and Neutralization of Antivenom
Abstract
:1. Introduction
2. Results and Discussion
2.1. Proteome of Trimeresurus nebularis Venom
2.2. Immunological Profiling of T. nebularis Venom
2.3. Cross-Neutralization of T. nebularis Venom Toxicity
3. Conclusions
4. Materials and Methods
4.1. Venoms and Antivenoms
4.2. Estimation of Antivenom Protein Concentration
4.3. Whole Venom In-Solution Tryptic Digestion and Protein Identification by Tandem Mass Spectrometry (Nano-ESI-LC-MS/MS)
4.4. Sodium Dodecyl Sulphate-Polyacrylamide Gel Electrophoresis (SDS-PAGE)
4.5. Immunological Binding Assay
4.6. Neutralization of T. nebularis Venom Procoagulant Effect
4.7. Neutralization of T. nebularis Venom Toxicity
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Protein Family/Protein Identity a | Database Accession b | Species c | Relative Abundance d | |
---|---|---|---|---|
Snake venom metalloproteinases (SVMP) | 48.42% | |||
P-II subtypes (1-8) | 15.92% | |||
1 | Zinc metalloproteinase/disintegrin | Q805F4 | Agkistrodon piscivorus piscivorus | 4.28% |
2 | Zinc metalloproteinase-disintegrin stejnihagin-B | CL4568.contig1_Cp | Trimeresurus purpureomaculatus | 3.20% |
3 | Zinc metalloproteinase-disintegrin stejnitin | P0DM87 | Trimeresurus stejnegeri | 2.60% |
4 | Zinc metalloproteinase/disintegrin | P0C6E4 | Protobothrops jerdonii | 2.53% |
5 | Metalloproteinase 3 | CL174.contig3_CrT | Calloselasma rhodostoma | 2.17% |
6 | metalloproteinase 9 | CL92.contig6_Ta | Trimeresurus albolabris | 0.74% |
7 | P-II metalloprotease | T2HRS1 | Protobothrops flavoviridis | 0.25% |
8 | Zinc metalloproteinase homolog-disintegrin albolatin | P0C6B6 | Trimeresurus albolabris | 0.15% |
P-III subtypes (9-19) | 32.50% | |||
9 | metalloproteinase isoform 1 | CL83.contig1_Ta | Trimeresurus albolabris | 6.63% |
10 | Zinc metalloproteinase-disintegrin ACLD | CL92.contig4_Ta | Trimeresurus albolabris | 5.94% |
11 | Zinc metalloproteinase-disintegrin TSV-DM | CL83.contig2_Ta | Trimeresurus albolabris | 3.06% |
12 | Zinc metalloproteinase-disintegrin-like stejnihagin-B | Q3HTN2 | Trimeresurus stejnegeri | 2.82% |
13 | Zinc metalloproteinase-disintegrin HV1 | Unigene635_Cp | Trimeresurus purpureomaculatus | 2.81% |
14 | group III snake venom metalloproteinase | E9KJZ5 | Echis ocellatus | 2.80% |
15 | Zinc metalloproteinase-disintegrin-like TSV-DM | Q2LD49 | Trimeresurus stejnegeri | 2.35% |
16 | Zinc metalloproteinase homolog/disintegrin | CL288.contig3_Ta | Trimeresurus albolabris | 2.24% |
17 | Zinc metalloproteinase-disintegrin stejnihagin-A | CL92.contig5_Ta | Trimeresurus albolabris | 1.67% |
18 | Snake venom metalloproteinase 5 | J3S831 | Crotalus adamanteus | 1.19% |
19 | Zinc metalloproteinase-disintegrin-like batroxstatin-3 | C5H5D4 | Bothrops atrox | 0.99% |
Snaclecs | 19.43% | |||
1 | Snaclec stejaggregin-B subunit beta-1 | Q71RQ9 | Trimeresurus stejnegeri | 5.03% |
2 | Snaclec coagulation factor IX/factor X-binding protein subunit A | Q71RR4 | Trimeresurus stejnegeri | 4.90% |
3 | Mucrocetin subunit alpha | Unigene86_Ta | Trimeresurus albolabris | 3.13% |
4 | Snaclec stejaggregin-B subunit alpha | Q71RQ7 | Trimeresurus stejnegeri | 2.88% |
5 | C-type lectin TsL | Q9YGP1 | Trimeresurus stejnegeri | 2.29% |
6 | C-type lectin Cal | P21963 | Crotalus atrox | 1.19% |
Snake venom serine proteases (SVSP) | 14.27% | |||
1 | Alpha-fibrinogenase albofibrase | P0CJ41 | Trimeresurus albolabris | 5.32% |
2 | Thrombin-like enzyme ancrod | P26324 | Calloselasma rhodostoma | 2.83% |
3 | Thrombin-like enzyme halystase | P81176 | Gloydius blomhoffii | 2.35% |
4 | Snake venom serine protease serpentokallikrein-2 | Q9DG84 | Protobothrops mucrosquamatus | 2.13% |
5 | Snake venom serine protease KN13 | Q71QH6 | Trimeresurus stejnegeri | 1.64% |
Phospholipases A2 | 5.40% | |||
1 | Phospholipase A2 | A0A0H3U232 | Trimeresurus sabahi | 4.27% |
2 | Basic phospholipase A2 homolog | P22640 | Protobothrops mucrosquamatus | 1.13% |
Disintegrins | 5.26% | |||
1 | Disintegrin ussuristatin-1 | Q7LZI5 | Gloydius ussuriensis | 2.67% |
2 | Disintegrin trigramin-gamma | P62383 | Trimeresurus gramineus | 2.59% |
Cysteine-rich secretory proteins (CRiSP) | 4.31% | |||
1 | Cysteine-rich venom protein | P60623 | Trimeresurus stejnegeri | 3.84% |
2 | Cysteine rich secretory protein | T2HP25 | Protobothrops flavoviridis | 0.47% |
L-amino acid oxidase (LAAO) | 0.99% | |||
1 | L-amino acid oxidase | CL43.contig1_Ta | Trimeresurus albolabris | 0.99% |
Phosphodiesterases (PDE) | 0.37% | |||
1 | Phosphodiesterase 1 | CL2883.contig1_Cp | Trimeresuruspurpureomaculatus | 0.29% |
2 | Venom phosphodiesterase 1 | J3SEZ3 | Crotalus adamanteus | 0.08% |
5′-nucleotidase (5′NUC) | 0.15% | |||
1 | Snake venom 5′-nucleotidase | CL554.contig1_Ta | Trimeresurus albolabris | 0.15% |
Cellular proteins | 1.42% | |||
1 | Endonuclease domain-containing 1 protein-like | Unigene20352_Ec | Echis carinatus | 0.63% |
2 | Endonuclease domain-containing 1 protein-like | Unigene20352_Ec | Echis carinatus | 0.41% |
3 | Endonuclease domain-containing 1 protein-like CPACP | CL3153.contig1_Cp | Trimeresuruspurpureomaculatus | 0.29% |
4 | Glutaminyl-peptide cyclotransferase | Q90YA8 | Gloydius blomhoffii | 0.08% |
Lethality | i.v. LD50 a (µg/g) | Challenge Dose | ED50 b (µL) | ER50 c (mg/mL) | Potency, P d (mg/mL) | Normalized Potency, n-P e (mg/g) |
2.00 (1.61–2.48) | 5 LD50 | 100.00 | 2.00 (1.61–2.48) | 1.6 | 79.2 | |
Procoagulant activity | MCD f (µg/mL) | Challenge Dose | ED g (µL) | ER h (mg/mL) | ||
150.0 ± 6.0 | 2 MCD | 13.2 ± 0.5 | 4.6 ± 0.2 | NA | NA | |
Hemorrhagic activity | MHD i (µg) | Challenge Dose | ED50 j (µL) | ER50 k (mg/mL) | ||
1.67 ± 0.15 | 2 MHD | 0.95 ± 0.13 | 3.52 ± 0.3 | NA | NA |
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Tan, C.H.; Tan, K.Y.; Ng, T.S.; Quah, E.S.H.; Ismail, A.K.; Khomvilai, S.; Sitprija, V.; Tan, N.H. Venomics of Trimeresurus (Popeia) nebularis, the Cameron Highlands Pit Viper from Malaysia: Insights into Venom Proteome, Toxicity and Neutralization of Antivenom. Toxins 2019, 11, 95. https://doi.org/10.3390/toxins11020095
Tan CH, Tan KY, Ng TS, Quah ESH, Ismail AK, Khomvilai S, Sitprija V, Tan NH. Venomics of Trimeresurus (Popeia) nebularis, the Cameron Highlands Pit Viper from Malaysia: Insights into Venom Proteome, Toxicity and Neutralization of Antivenom. Toxins. 2019; 11(2):95. https://doi.org/10.3390/toxins11020095
Chicago/Turabian StyleTan, Choo Hock, Kae Yi Tan, Tzu Shan Ng, Evan S.H. Quah, Ahmad Khaldun Ismail, Sumana Khomvilai, Visith Sitprija, and Nget Hong Tan. 2019. "Venomics of Trimeresurus (Popeia) nebularis, the Cameron Highlands Pit Viper from Malaysia: Insights into Venom Proteome, Toxicity and Neutralization of Antivenom" Toxins 11, no. 2: 95. https://doi.org/10.3390/toxins11020095