Rapid and Efficient Enrichment of Snake Venoms from Human Plasma Using a Strong Cation Exchange Tip Column to Improve Snakebite Diagnosis
Abstract
:1. Introduction
2. Results
2.1. Differences in pI Values between High-Abundance Human Plasma Proteins and Snake Venom Proteins
2.2. SCX-HPLC for Separating Venom Proteins from Human Plasma Proteins
2.3. Development of Methodology for Efficient Enrichment of Snake Venom Proteins from Human Plasma
2.4. Identification of SCX Tip Column-Enriched Snake Venom Proteins
2.5. Evaluation of Venom Protein Enrichment Efficiency Via the SCX Tip Column
2.6. Combination of SCX-Tip Column Enrichment with Snakebite Detection Assays
3. Discussion
4. Conclusions
5. Materials and Methods
5.1. Snake Venom
5.2. Strong Cation Exchange High-Performance Liquid Chromatography (SCX-HPLC)
5.3. Separation of Venom and Plasma Proteins Using the Strong Cation Exchange (SCX) Tip Column
5.4. Acetone Precipitation
5.5. In-Gel Tryptic Digestion
5.6. LC-MS/MS Analysis
5.7. Database Searches and Bioinformatics Analysis
5.8. Venom Detection with Lateral Flow Strips
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Acknowledgments
Conflicts of Interest
References
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Species | Sample | Accession | Description | Coverage (%) 1 | Peptides 2 | PSMs 3 | MW [kDa] | pI |
---|---|---|---|---|---|---|---|---|
N. atra | 100mM-1 | P60304 | Cytotoxin 1 | 35.8 | 4 | 10 | 9.0 | 8.7 |
100mM-2 | P60304 | Cytotoxin 1 | 71.6 | 11 | 229 | 9.0 | 8.7 | |
P80245 | Cytotoxin 6 | 58.0 | 8 | 103 | 9.0 | 8.9 | ||
P01442 | Cytotoxin 2 | 51.9 | 7 | 86 | 9.0 | 8.9 | ||
P01443 | Cytotoxin 4 | 51.9 | 7 | 86 | 9.1 | 9.0 | ||
150 mM-1 | P60304 | Cytotoxin 1 | 35.8 | 5 | 12 | 9.0 | 8.7 | |
P80245 | Cytotoxin 6 | 43.2 | 5 | 10 | 9.0 | 8.9 | ||
150 mM-2 | O93422 | Long neurotoxin homolog | 22.1 | 2 | 6 | 9.8 | 8.7 | |
Q9YGI2 | Probable weak neurotoxin NNAM1 | 22.1 | 2 | 6 | 9.8 | 8.5 | ||
Q9YGI4 | Probable weak neurotoxin NNAM2 | 25.6 | 2 | 10 | 9.9 | 8.6 | ||
150 mM-3 | P60304 | Cytotoxin 1 | 71.6 | 11 | 356 | 9.0 | 8.7 | |
P80245 | Cytotoxin 6 | 71.6 | 12 | 164 | 9.0 | 8.9 | ||
P01442 | Cytotoxin 2 | 59.3 | 9 | 124 | 9.0 | 8.9 | ||
P01443 | Cytotoxin 4 | 59.3 | 9 | 124 | 9.1 | 9.0 | ||
200 mM-1 | Q7ZZN8 | Cysteine-rich venom protein natrin-2 | 33.6 | 9 | 63 | 26.2 | 8.7 | |
200 mM-2 | Q9YGI2 | Probable weak neurotoxin NNAM1 | 36.1 | 4 | 99 | 9.8 | 8.5 | |
O93422 | Long neurotoxin homolog | 36.1 | 4 | 99 | 9.8 | 8.7 | ||
Q9YGI4 | Probable weak neurotoxin NNAM2 | 25.6 | 2 | 37 | 9.9 | 8.6 | ||
200 mM-3 | P60301 | Cytotoxin 3 | 60.5 | 9 | 178 | 9.0 | 9.0 | |
P60304 | Cytotoxin 1 | 65.4 | 9 | 161 | 9.0 | 8.7 | ||
P80245 | Cytotoxin 6 | 58.0 | 9 | 115 | 9.0 | 8.9 | ||
250 mM-1 | P62375 | Cytotoxin A5 | 45.8 | 4 | 44 | 9.3 | 9.0 | |
300 mM-1 | P62375 | Cytotoxin A5 | 46.0 | 6 | 17 | 9.3 | 9.0 | |
B. multicinctus | 150 mM-1 | P00617 | Basic phospholipase A2 beta-bungarotoxin A1 chain | 21.1 | 2 | 5 | 16.2 | 7.5 |
150 mM-2 | P00987 | Kunitz-type serine protease inhibitor homolog beta-bungarotoxin B1 chain | 28.2 | 3 | 17 | 9.6 | 8.7 | |
200 mM-1 | P00617 | Basic phospholipase A2 beta-bungarotoxin A1 chain | 61.9 | 9 | 44 | 16.2 | 7.5 | |
200 mM-2 | P00987 | Kunitz-type serine protease inhibitor homolog beta-bungarotoxin B1 chain | 48.2 | 5 | 26 | 9.6 | 8.7 | |
250 mM-1 | P00617 | Basic phospholipase A2 beta-bungarotoxin A1 chain | 53.1 | 7 | 17 | 16.2 | 7.5 | |
250 mM-2 | P00987 | Kunitz-type serine protease inhibitor homolog beta-bungarotoxin B1 chain | 30.6 | 3 | 9 | 9.6 | 8.7 |
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Liu, C.-C.; Yang, Y.-H.; Hsiao, Y.-C.; Wang, P.-J.; Liu, J.-C.; Liu, C.-H.; Hsieh, W.-C.; Lin, C.-C.; Yu, J.-S. Rapid and Efficient Enrichment of Snake Venoms from Human Plasma Using a Strong Cation Exchange Tip Column to Improve Snakebite Diagnosis. Toxins 2021, 13, 140. https://doi.org/10.3390/toxins13020140
Liu C-C, Yang Y-H, Hsiao Y-C, Wang P-J, Liu J-C, Liu C-H, Hsieh W-C, Lin C-C, Yu J-S. Rapid and Efficient Enrichment of Snake Venoms from Human Plasma Using a Strong Cation Exchange Tip Column to Improve Snakebite Diagnosis. Toxins. 2021; 13(2):140. https://doi.org/10.3390/toxins13020140
Chicago/Turabian StyleLiu, Chien-Chun, Ya-Han Yang, Yung-Chin Hsiao, Po-Jung Wang, Jo-Chuan Liu, Chien-Hsin Liu, Wen-Chin Hsieh, Chih-Chuan Lin, and Jau-Song Yu. 2021. "Rapid and Efficient Enrichment of Snake Venoms from Human Plasma Using a Strong Cation Exchange Tip Column to Improve Snakebite Diagnosis" Toxins 13, no. 2: 140. https://doi.org/10.3390/toxins13020140
APA StyleLiu, C.-C., Yang, Y.-H., Hsiao, Y.-C., Wang, P.-J., Liu, J.-C., Liu, C.-H., Hsieh, W.-C., Lin, C.-C., & Yu, J.-S. (2021). Rapid and Efficient Enrichment of Snake Venoms from Human Plasma Using a Strong Cation Exchange Tip Column to Improve Snakebite Diagnosis. Toxins, 13(2), 140. https://doi.org/10.3390/toxins13020140