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dc.creatorSilva Soares, Bárbara da
dc.creatorRocha, Surza Lucia Gonçalves
dc.creatorAlmeida Bastos, Viviane
dc.creatorBorges Lima, Diogo
dc.creatorCosta Carvalho, Paulo
dc.creatorGozzo, Fabio Cesar
dc.creatorDemeler, Borries
dc.creatorWilliams, Tayler L.
dc.creatorArnold, Janelle
dc.creatorHenrickson, Amy
dc.creatorJørgensen, Thomas J.D.
dc.creatorSouza, ‪Tatiana de Arruda Campos Brasil de
dc.creatorPerales, Jonas
dc.creatorValente, Richard H.
dc.creatorLomonte, Bruno
dc.creatorGomes Neto, Francisco
dc.creatorNeves Ferreira, Ana Gisele da Costa
dc.date.accessioned2022-02-23T14:45:22Z
dc.date.available2022-02-23T14:45:22Z
dc.date.issued2022
dc.identifier.citationhttps://www.frontiersin.org/articles/10.3389/fmolb.2021.787368/full
dc.identifier.issn2296-889X
dc.identifier.urihttps://hdl.handle.net/10669/85839
dc.description.abstractDM64 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.es_ES
dc.description.sponsorshipFundação Oswaldo Cruz/[INOVA GC VPPCB-007-FIO-18-2-9]/Fiocruz/Brasiles_ES
dc.description.sponsorshipFundação de Amparo à Pesquisa do Estado do Rio de Janeiro/[APQ1 E-6/010.001929/2019]/FAPERJ/Brasiles_ES
dc.description.sponsorshipConselho Nacional de Desenvolvimento Científico e Tecnológico/[Universal 426290/2018-6]/CNPq/Brasiles_ES
dc.description.sponsorshipNational Institutes of Health/[GM120600]/NIH/Estados Unidoses_ES
dc.description.sponsorshipNational Science Foundation/[NSF-ACI-1339649]/NSF/Estados Unidoses_ES
dc.description.sponsorshipSan Diego Supercomputer Center/[TG-MCB070039N]/SDSC/Estados Unidoses_ES
dc.description.sponsorshipTexas Advanced Computing Center/[TG457201]/TACC/Estados Unidoses_ES
dc.language.isoenges_ES
dc.sourceFrontiers in Molecular Biosciences,, vol.8, pp.1-24.es_ES
dc.subjectCross-linking (XL)es_ES
dc.subjectMass spectrometryes_ES
dc.subjectImmunoglobulin foldes_ES
dc.subjectStructural biologyes_ES
dc.subjectToxin neutralisationes_ES
dc.subjectProtein inhibitores_ES
dc.subjectSnake envenomationes_ES
dc.subjectAntiophidic activityes_ES
dc.subjectVeneno de serpientees_ES
dc.titleMolecular architecture of the antiophidic protein DM64 and its binding specificity to myotoxin II from Bothrops asper venomes_ES
dc.typeartículo científico
dc.identifier.doi10.3389/fmolb.2021.787368
dc.description.procedenceUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP)es_ES


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