{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,30]],"date-time":"2026-09-30T20:09:35Z","timestamp":1790798975986,"version":"4.1.0"},"reference-count":57,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2023,9,5]],"date-time":"2023-09-05T00:00:00Z","timestamp":1693872000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Wellcome Trust","award":["221710\/Z\/20\/Z"],"award-info":[{"award-number":["221710\/Z\/20\/Z"]}]},{"name":"Wellcome Trust","award":["221712\/Z\/20\/Z"],"award-info":[{"award-number":["221712\/Z\/20\/Z"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Toxins"],"abstract":"<jats:p>Modern analytical size exclusion chromatography (SEC) is a suitable technique to separate venom toxin families according to their size characteristics. In this study, a method was developed to separate intact venom toxins from Bungarus multicinctus and Daboia russelii venoms via analytical SEC using volatile, non-salt-containing eluents for post-column mass spectrometry, coagulation bioassaying and high-throughput venomics. Two venoms were used to demonstrate the method developed. While the venom of Bungaurs multicinctus is known to exert anticoagulant effects on plasma, in this study, we showed the existence of both procoagulant toxins and anticoagulant toxins. For Daboia russelii venom, the method revealed characteristic procoagulant effects, with a 90 kDa mass toxin detected and matched with the Factor X-activating procoagulant heterotrimeric glycoprotein named RVV-X. The strong procoagulant effects for this toxin show that it was most likely eluted from size exclusion chromatography non-denatured. In conclusion, the separation of snake venom by size gave the opportunity to separate some specific toxin families from each other non-denatured, test these for functional bioactivities, detect the eluting mass on-line via mass spectrometry and identify the eluted toxins using high-throughput venomics.<\/jats:p>","DOI":"10.3390\/toxins15090552","type":"journal-article","created":{"date-parts":[[2023,9,5]],"date-time":"2023-09-05T10:26:43Z","timestamp":1693909603000},"page":"552","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Analytical Size Exclusion Chromatography Coupled with Mass Spectrometry in Parallel with High-Throughput Venomics and Bioassaying for Venom Profiling"],"prefix":"10.3390","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0009-0000-0598-2240","authenticated-orcid":false,"given":"Sedef","family":"Terzioglu","sequence":"first","affiliation":[{"name":"Division of Bioanalytical Chemistry, Department of Chemistry and Pharmaceutical Sciences, Faculty of Sciences, Amsterdam Institute of Molecular and Life Sciences (AIMMS), Vrije Universiteit Amsterdam, 1081 HZ Amsterdam, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"M\u00e1ty\u00e1s A.","family":"Bittenbinder","sequence":"additional","affiliation":[{"name":"Division of Bioanalytical Chemistry, Department of Chemistry and Pharmaceutical Sciences, Faculty of Sciences, Amsterdam Institute of Molecular and Life Sciences (AIMMS), Vrije Universiteit Amsterdam, 1081 HZ Amsterdam, The Netherlands"},{"name":"Naturalis Biodiversity Center, 2333 CR Leiden, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Julien","family":"Slagboom","sequence":"additional","affiliation":[{"name":"Division of Bioanalytical Chemistry, Department of Chemistry and Pharmaceutical Sciences, Faculty of Sciences, Amsterdam Institute of Molecular and Life Sciences (AIMMS), Vrije Universiteit Amsterdam, 1081 HZ Amsterdam, The Netherlands"},{"name":"Centre for Analytical Sciences Amsterdam (CASA), 1098 XH Amsterdam, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bas","family":"van de Velde","sequence":"additional","affiliation":[{"name":"Division of Bioanalytical Chemistry, Department of Chemistry and Pharmaceutical Sciences, Faculty of Sciences, Amsterdam Institute of Molecular and Life Sciences (AIMMS), Vrije Universiteit Amsterdam, 1081 HZ Amsterdam, The Netherlands"},{"name":"Centre for Analytical Sciences Amsterdam (CASA), 1098 XH Amsterdam, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8035-4719","authenticated-orcid":false,"given":"Nicholas R.","family":"Casewell","sequence":"additional","affiliation":[{"name":"Centre for Snakebite Research & Interventions, Liverpool School of Tropical Medicine, Liverpool L3 5QA, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0011-5612","authenticated-orcid":false,"given":"Jeroen","family":"Kool","sequence":"additional","affiliation":[{"name":"Division of Bioanalytical Chemistry, Department of Chemistry and Pharmaceutical Sciences, Faculty of Sciences, Amsterdam Institute of Molecular and Life Sciences (AIMMS), Vrije Universiteit Amsterdam, 1081 HZ Amsterdam, The Netherlands"},{"name":"Centre for Analytical Sciences Amsterdam (CASA), 1098 XH Amsterdam, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,9,5]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"17063","DOI":"10.1038\/nrdp.2017.63","article-title":"Snakebite Envenoming","volume":"3","author":"Calvete","year":"2017","journal-title":"Nat. Rev. Dis. Prim."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1736","DOI":"10.1016\/j.febslet.2009.03.029","article-title":"Venoms, Venomics, Antivenomics","volume":"583","author":"Calvete","year":"2009","journal-title":"FEBS Lett."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1081\/TXR-120019018","article-title":"Effectiveness of Snake Antivenom: Species and Regional Venom Variation and Its Clinical Impact","volume":"22","author":"Fry","year":"2003","journal-title":"J. Toxicol. Toxin Rev."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"570","DOI":"10.1016\/j.tips.2020.05.006","article-title":"Causes and Consequences of Snake Venom Variation","volume":"41","author":"Casewell","year":"2020","journal-title":"Trends Pharmacol. Sci."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"109063","DOI":"10.1016\/j.cbpc.2021.109063","article-title":"Proteomics and Neutralization of Bungarus multicinctus (Many-Banded Krait) Venom: Intra-Specific Comparisons between Specimens from China and Taiwan","volume":"247","author":"Oh","year":"2021","journal-title":"Comp. Biochem. Physiol. Part C Toxicol. Pharmacol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1735","DOI":"10.1016\/j.jprot.2011.05.027","article-title":"Ending the Drought: New Strategies for Improving the Flow of Affordable, Effective Antivenoms in Asia and Africa","volume":"74","author":"Williams","year":"2011","journal-title":"J. Proteomics"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"827","DOI":"10.1016\/j.toxicon.2003.11.002","article-title":"Excitement Ahead: Structure, Function and Mechanism of Snake Venom Phospholipase A2 Enzymes","volume":"42","author":"Kini","year":"2003","journal-title":"Toxicon"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"114105","DOI":"10.1016\/j.bcp.2020.114105","article-title":"Snake Venom Three-Finger Toxins and Their Potential in Drug Development Targeting Cardiovascular Diseases","volume":"181","author":"Kini","year":"2020","journal-title":"Biochem. Pharmacol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"3","DOI":"10.1016\/j.toxicon.2012.09.004","article-title":"Snake Venom Metalloproteinases","volume":"62","author":"Markland","year":"2013","journal-title":"Toxicon"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"19","DOI":"10.1016\/j.toxicon.2012.09.003","article-title":"The Long Road of Research on Snake Venom Serine Proteinases","volume":"62","author":"Serrano","year":"2013","journal-title":"Toxicon"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"4544","DOI":"10.1111\/j.1742-4658.2011.08115.x","article-title":"Enzymatic Toxins from Snake Venom: Structural Characterization and Mechanism of Catalysis","volume":"278","author":"Kang","year":"2011","journal-title":"FEBS J."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1405","DOI":"10.1002\/jms.1242","article-title":"Snake Venomics. Strategy and Applications","volume":"42","author":"Calvete","year":"2007","journal-title":"J. Mass Spectrom."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"135","DOI":"10.1016\/S0021-9673(99)00546-4","article-title":"How Does a Protein Unfold on a Reversed-Phase Liquid Chromatography Surface?","volume":"849","author":"McNay","year":"1999","journal-title":"J. Chromatogr. A"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"195","DOI":"10.1016\/S0167-4838(98)00121-6","article-title":"Acetonitrile-Protein Interactions: Amino Acid Solubility and Preferential Solvation","volume":"1387","author":"Gekko","year":"1998","journal-title":"Biochim. Biophys. Acta Protein Struct. Mol. Enzymol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1101","DOI":"10.1110\/ps.062323407","article-title":"Molecular Dynamics Simulations of the Native and Partially Folded States of Ubiquitin: Influence of Methanol Cosolvent, PH, and Temperature on the Protein Structure and Dynamics","volume":"16","author":"Kony","year":"2007","journal-title":"Protein Sci."},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Yee, K.T., Pitts, M., Tongyoo, P., Rojnuckarin, P., and Wilkinson, M.C. (2017). Snake Venom Metalloproteinases and Their Peptide Inhibitors from Myanmar Russell\u2019s Viper Venom. Toxins, 9.","DOI":"10.3390\/toxins9010015"},{"key":"ref_17","first-page":"1646","article-title":"Effects of Sodium Chloride Concentration on Growth, Biochemical Composition, and Metabolism of HeLa Cells","volume":"20","author":"Stubblefield","year":"1960","journal-title":"Cancer Res."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1045","DOI":"10.1016\/j.jasms.2010.02.003","article-title":"Effects of Buffer Loading for Electrospray Ionization Mass Spectrometry of a Noncovalent Protein Complex That Requires High Concentrations of Essential Salts","volume":"21","author":"Sterling","year":"2010","journal-title":"J. Am. Soc. Mass Spectrom."},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Slagboom, J., Mladi\u0107, M., Xie, C., Kazandjian, T.D., Vonk, F., Somsen, G.W., Casewell, N.R., and Kool, J. (2020). High Throughput Screening and Identification of Coagulopathic Snake Venom Proteins and Peptides Using Nanofractionation and Proteomics Approaches. PLoS Negl. Trop. Dis., 14.","DOI":"10.1371\/journal.pntd.0007802"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"2594","DOI":"10.1002\/elps.201500207","article-title":"Fractionation and Proteomic Analysis of the Walterinnesia Aegyptia Snake Venom Using OFFGEL and MALDI-TOF-MS Techniques","volume":"36","author":"Aziz","year":"2015","journal-title":"Electrophoresis"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Slagboom, J., Derks, R.J.E., Sadighi, R., Somsen, G.W., Ulens, C., Casewell, N.R., and Kool, J. (2023). High-Throughput Venomics. J. Proteome Res.","DOI":"10.1021\/acs.jproteome.2c00780"},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Still, K.B.M., Nandlal, R.S.S., Slagboom, J., Somsen, G.W., Casewell, N.R., and Kool, J. (2017). Multipurpose HTS Coagulation Analysis: Assay Development and Assessment of Coagulopathic Snake Venoms. Toxins, 9.","DOI":"10.3390\/toxins9120382"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.jprot.2018.05.003","article-title":"Proteomics, Functional Characterization and Antivenom Neutralization of the Venom of Pakistani Russell\u2019s Viper (Daboia russelii) from the Wild","volume":"183","author":"Faisal","year":"2018","journal-title":"J. Proteomics"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1186\/1471-2164-12-1","article-title":"Venom Gland Transcriptomes of Two Elapid Snakes (Bungarus multicinctus and Naja Atra) and Evolution of Toxin Genes","volume":"12","author":"Jiang","year":"2011","journal-title":"BMC Genom."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"368","DOI":"10.1016\/j.jchromb.2018.06.029","article-title":"Unraveling the Mysteries of Modern Size Exclusion Chromatography\u2014The Way to Achieve Confident Characterization of Therapeutic Proteins","volume":"1092","author":"Goyon","year":"2018","journal-title":"J. Chromatogr. B Anal. Technol. Biomed. Life Sci."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"80","DOI":"10.1016\/j.chroma.2016.11.056","article-title":"Evaluation of Size Exclusion Chromatography Columns Packed with Sub-3 \u039cm Particles for the Analysis of Biopharmaceutical Proteins","volume":"1498","author":"Goyon","year":"2017","journal-title":"J. Chromatogr. A"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"73","DOI":"10.1016\/S0021-9673(97)00343-9","article-title":"Influence of Differences between Sample and Mobile Phase Viscosities on the Shape of Chromatographic Elution Profiles","volume":"775","author":"Castells","year":"1997","journal-title":"J. Chromatogr. A"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"100733","DOI":"10.1016\/j.blre.2020.100733","article-title":"Interplay between Platelets and Coagulation","volume":"46","author":"Sang","year":"2021","journal-title":"Blood Rev."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"53","DOI":"10.1134\/S1607672915010159","article-title":"Venoms of Kraits Bungarus multicinctus and Bungarus fasciatus Contain Anticoagulant Proteins","volume":"460","author":"Utkin","year":"2015","journal-title":"Dokl. Biochem. Biophys."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"379","DOI":"10.1097\/MBC.0000000000000842","article-title":"Anticoagulant Activity of Krait, Coral Snake, and Cobra Neurotoxic Venoms with Diverse Proteomes Are Inhibited by Carbon Monoxide","volume":"30","author":"Nielsen","year":"2019","journal-title":"Blood Coagul. Fibrinolysis"},{"key":"ref_31","first-page":"E80","article-title":"Local Tissue Necrosis and Thrombocytopenia Following Bungarus multicinctus Envenomation in a Child","volume":"119","author":"Wang","year":"2021","journal-title":"Arch. Argent. Pediatr."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"369","DOI":"10.1016\/j.peptides.2007.11.013","article-title":"A Novel Serine Protease Inhibitor from Bungarus fasciatus Venom","volume":"29","author":"Lu","year":"2008","journal-title":"Peptides"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"147","DOI":"10.1590\/S1678-91992010005000009","article-title":"Enzymatic and Immunological Properties of Bungarus flaviceps (Red-Headed Krait) Venom","volume":"16","author":"Tan","year":"2010","journal-title":"J. Venom. Anim. Toxins Incl. Trop. Dis."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"1277","DOI":"10.1016\/0041-0101(95)00070-3","article-title":"An Activator of Blood Coagulation Factor X from the Venom of Bungarus fasciatus","volume":"33","author":"Zhang","year":"1995","journal-title":"Toxicon"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"197","DOI":"10.1016\/j.toxicon.2015.08.026","article-title":"Quantitative Proteomic Analysis of Vietnamese Krait Venoms: Neurotoxins Are the Major Components in Bungarus multicinctus and Phospholipases A2 in Bungarus fasciatus","volume":"107","author":"Ziganshin","year":"2015","journal-title":"Toxicon"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1016\/j.jprot.2016.02.028","article-title":"Proteomic Characterization and Comparison of Venoms from Two Elapid Snakes (Bungarus multicinctus and Naja Atra) from China","volume":"138","author":"Shan","year":"2016","journal-title":"J. Proteomics"},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Xie, C., Albulescu, L.-O., Bittenbinder, M.A., Somsen, G.W., Vonk, F.J., Casewell, N.R., and Kool, J. (2020). Neutralizing Effects of Small Molecule Inhibitors and Metal Chelators on Coagulopathic Viperinae Snake Venom Toxins. Biomedicines, 8.","DOI":"10.1101\/2020.06.02.129643"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"1835","DOI":"10.1016\/j.apsb.2020.09.005","article-title":"Neutralising Effects of Small Molecule Toxin Inhibitors on Nanofractionated Coagulopathic Crotalinae Snake Venoms","volume":"10","author":"Xie","year":"2020","journal-title":"Acta Pharm. Sin. B"},{"key":"ref_39","first-page":"1","article-title":"Proteomics, Toxicity and Antivenom Neutralization of Sri Lankan and Indian Russell\u2019s Viper (Daboia russelii) Venoms","volume":"27","author":"Faisal","year":"2021","journal-title":"J. Venom. Anim. Toxins Incl. Trop. Dis."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"153","DOI":"10.1016\/j.biochi.2013.11.026","article-title":"A Major Phospholipase A2 from Daboia russelii Russelii Venom Shows Potent Anticoagulant Action via Thrombin Inhibition and Binding with Plasma Phospholipids","volume":"99","author":"Mukherjee","year":"2014","journal-title":"Biochimie"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"291","DOI":"10.1016\/j.toxicon.2013.10.001","article-title":"Mechanism of in Vivo Anticoagulant and Haemolytic Activity by a Neutral Phospholipase A2 Purified from Daboia russelii Russelii Venom: Correlation with Clinical Manifestations in Russell\u2019s Viper Envenomed Patients","volume":"76","author":"Saikia","year":"2013","journal-title":"Toxicon"},{"key":"ref_42","doi-asserted-by":"crossref","unstructured":"Sharma, M., Iyer, J.K., Shih, N., Majumder, M., Mattaparthi, V.S.K., Mukhopadhyay, R., and Doley, R. (2016). Daboxin p, a Major Phospholipase A2 Enzyme from the Indian Daboia russelii Russelii Venom Targets Factor x and Factor Xa for Its Anticoagulant Activity. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0153770"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"1150","DOI":"10.1016\/j.thromres.2014.09.009","article-title":"A New C-Type Lectin (RVsnaclec) Purified from Venom of Daboia russelii Russelii Shows Anticoagulant Activity via Inhibition of FXa and Concentration-Dependent Differential Response to Platelets in a Ca2+-Independent Manner","volume":"134","author":"Mukherjee","year":"2014","journal-title":"Thromb. Res."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"154","DOI":"10.1016\/j.ijbiomac.2014.02.058","article-title":"Characterization of a Kunitz-Type Protease Inhibitor Peptide (Rusvikunin) Purified from Daboia russelii Russelii Venom","volume":"67","author":"Mukherjee","year":"2014","journal-title":"Int. J. Biol. Macromol."},{"key":"ref_45","doi-asserted-by":"crossref","unstructured":"Tasoulis, T., and Isbister, G.K. (2017). A Review and Database of Snake Venom Proteomes. Toxins, 9.","DOI":"10.3390\/toxins9090290"},{"key":"ref_46","doi-asserted-by":"crossref","unstructured":"Lin, B., Zhang, J.R., Lu, H.J., Zhao, L., Chen, J., Zhang, H.F., Wei, X.S., Zhang, L.Y., Wu, X.B., and Lee, W.H. (2020). Immunoreactivity and Neutralization Study of Chinese Bungarus multicinctus Antivenin and Lab-Prepared Anti-Bungarotoxin Antisera towards Purified Bungarotoxins and Snake Venoms. PLoS Negl. Trop. Dis., 14.","DOI":"10.1371\/journal.pntd.0008873"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"2","DOI":"10.1371\/journal.pone.0105404","article-title":"Recognition of Bungarus multicinctus Venom by a DNA Aptamer against \u03b2-Bungarotoxin","volume":"9","author":"Ye","year":"2014","journal-title":"PLoS ONE"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"338347","DOI":"10.1016\/j.aca.2021.338347","article-title":"Alternative Mobile Phase Additives for the Characterization of Protein Biopharmaceuticals in Liquid Chromatography\u2014Mass Spectrometry","volume":"1156","author":"Lardeux","year":"2021","journal-title":"Anal. Chim. Acta"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"10644","DOI":"10.1016\/S0021-9258(17)34108-X","article-title":"Factor X-Activating Glycoprotein of Russell\u2019s Viper Venom. Polypeptide Composition and Characterization of the Carbohydrate Moieties","volume":"269","author":"Gowda","year":"1994","journal-title":"J. Biol. Chem."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"3944","DOI":"10.1111\/j.1742-4658.2008.06540.x","article-title":"New Insights into the Functions and N-Glycan Structures of Factor X Activator from Russell\u2019s Viper Venom","volume":"275","author":"Chen","year":"2008","journal-title":"FEBS J."},{"key":"ref_51","doi-asserted-by":"crossref","unstructured":"Kini, R.M., Clemetson, K.J., Markland, F.S., McLane, M.A., and Morita, T. (2010). Toxins and Hemostasis: From Bench to Bedside, Springer.","DOI":"10.1007\/978-90-481-9295-3"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"5987","DOI":"10.1007\/s00216-017-0531-3","article-title":"Rapid Screening and Identification of ACE Inhibitors in Snake Venoms Using At-Line Nanofractionation LC-MS","volume":"409","author":"Mladic","year":"2017","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"141","DOI":"10.4155\/pbp.14.7","article-title":"A High Throughput Ultra Performance Size Exclusion Chromatography Assay for the Analysis of Aggregates and Fragments of Monoclonal Antibodies","volume":"2","author":"Mou","year":"2014","journal-title":"Pharm. Bioprocess."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"761","DOI":"10.1016\/S0959-440X(01)00278-0","article-title":"Proteins in Organic Solvents","volume":"11","author":"Mattos","year":"2001","journal-title":"Curr. Opin. Struct. Biol."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"1115","DOI":"10.1016\/j.toxicon.2005.02.020","article-title":"Snake Venom Serine Proteinases: Sequence Homology vs. Substrate Specificity, a Paradox to Be Solved","volume":"45","author":"Serrano","year":"2005","journal-title":"Toxicon"},{"key":"ref_56","first-page":"49","article-title":"Phospholipase A2 Biochemistry. Cardiovasc","volume":"23","author":"Burke","year":"2009","journal-title":"Drugs Ther."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"110","DOI":"10.1007\/s00239-003-2461-2","article-title":"Molecular Evolution and Phylogeny of Elapid Snake Venom Three-Finger Toxins","volume":"57","author":"Fry","year":"2003","journal-title":"J. Mol. Evol."}],"container-title":["Toxins"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2072-6651\/15\/9\/552\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T20:46:30Z","timestamp":1760129190000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2072-6651\/15\/9\/552"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,9,5]]},"references-count":57,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2023,9]]}},"alternative-id":["toxins15090552"],"URL":"https:\/\/doi.org\/10.3390\/toxins15090552","relation":{},"ISSN":["2072-6651"],"issn-type":[{"value":"2072-6651","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,9,5]]}}}