Show simple item record Soares, Barbara S. Rocha, Surza Lucia G. Bastos, Viviane A. Lima, Diogo B. Carvalho, Paulo C. Gozzo, Fabio C. Demeler, Borries Williams, Tayler L. Arnold, Janelle Henrickson, Amy Jorgensen, Thomas J. D. Souza, Tatiana A. C. B. Perales, Jonas Valente, Richard H. Lomonte, Bruno Gomes-Neto, Francisco Neves-Ferreira, Ana Gisele C. 2022-09-09T21:52:43Z 2022-09-09T21:52:43Z 2022
dc.identifier.citation Soares, B. S., Rocha, S. L. G., Bastos, V. A., Lima, D. B., Carvalho, P. C., Gozzo, F. C., Demeler, B., Williams, T. L., Arnold, J., Henrickson, A., Jorgensen, T. J. D., Souza, T. A. C. B., Perales, J., Valente, R. H., Lomonte, B., Gomes-Neto, F., & Neves-Ferreira, A. G. C. (2022). Molecular architecture of the antiophidic protein DM64 and its binding specificity to myotoxin II from Bothrops asper venom. Frontiers in Molecular Biosciences, 8, Article 787368. en_US
dc.description Open access article. Creative Commons Attribution 4.0 International License (CC BY 4.0) applies en_US
dc.description.abstract DM64 is a toxin-neutralizing serum glycoprotein isolated from Didelphis aurita, an ophiophagous marsupial naturally resistant to snake envenomation. This 64 kDa antitoxin targets myotoxic phospholipases A2, which account for most local tissue damage of viperid snakebites. We investigated the noncovalent complex formed between native DM64 and myotoxin II, a myotoxic phospholipase-like protein from Bothrops asper venom. Analytical ultracentrifugation (AUC) and size exclusion chromatography indicated that DM64 is monomeric in solution and binds equimolar amounts of the toxin. Attempts to crystallize native DM64 for X-ray diffraction were unsuccessful. Obtaining recombinant protein to pursue structural studies was also challenging. Classical molecular modeling techniques were impaired by the lack of templates with more than 25% sequence identity with DM64. An integrative structural biology approach was then applied to generate a three-dimensional model of the inhibitor bound to myotoxin II. I-TASSER individually modeled the five immunoglobulin-like domains of DM64. Distance constraints generated by cross-linking mass spectrometry of the complex guided the docking of DM64 domains to the crystal structure of myotoxin II, using Rosetta. AUC, small-angle X-ray scattering (SAXS), molecular modeling, and molecular dynamics simulations indicated that the DM64-myotoxin II complex is structured, shows flexibility, and has an anisotropic shape. Inter-protein cross-links and limited hydrolysis analyses shed light on the inhibitor’s regions involved with toxin interaction, revealing the critical participation of the first, third, and fifth domains of DM64. Our data showed that the fifth domain of DM64 binds to myotoxin II amino-terminal and beta-wing regions. The third domain of the inhibitor acts in a complementary way to the fifth domain. Their binding to these toxin regions presumably precludes dimerization, thus interfering with toxicity, which is related to the quaternary structure of the toxin. The first domain of DM64 interacts with the functional site of the toxin putatively associated with membrane anchorage. We propose that both mechanisms concur to inhibit myotoxin II toxicity by DM64 binding. The present topological characterization of this toxin-antitoxin complex constitutes an essential step toward the rational design of novel peptide-based antivenom therapies targeting snake venom myotoxins. en_US
dc.language.iso en_US en_US
dc.publisher Frontiers Research Foundation en_US
dc.subject Cross-linking (XL) en_US
dc.subject Immunogolubin fold en_US
dc.subject Structural biology en_US
dc.subject Toxin neutralisation en_US
dc.subject Protein inhibitor en_US
dc.subject Snake envenomation en_US
dc.subject Antiophidic activity en_US
dc.subject DM64
dc.subject.lcsh Proteins--Crosslinking
dc.subject.lcsh Mass spectrometry
dc.subject.lcsh Bothrops
dc.subject.lcsh Poisonous snakes--Venom
dc.subject.lcsh Snakebites
dc.title Molecular architecture of the antiophidic protein DM64 and its binding specificity to myotoxin II from Bothrops aasper venom en_US
dc.type Article en_US
dc.publisher.faculty Arts and Science en_US
dc.publisher.department Department of Chemistry and Biochemistry en_US
dc.description.peer-review Yes en_US
dc.publisher.institution Oswaldo Cruz Institute en_US
dc.publisher.institution Leibniz Forschungsinstitut für Molekulare Pharmakologie (FMP) en_US
dc.publisher.institution Carlos Chagas Institute en_US
dc.publisher.institution University of Campinas en_US
dc.publisher.institution University of Texas Health Science Center at San Antonio en_US
dc.publisher.institution University of Lethbridge en_US
dc.publisher.institution University of Montana en_US
dc.publisher.institution Princeton University en_US
dc.publisher.institution University of Southern Denmark en_US
dc.publisher.institution University of Costa Rica en_US
dc.publisher.url en_US

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