Please use this identifier to cite or link to this item: https://www.arca.fiocruz.br/handle/icict/31092
Title: Conformational plasticity of DM43, a metalloproteinase inhibitor from Didelphis marsupialis: chemical and pressure-induced equilibrium (un)folding studies
Authors: Chapeaurouge, Alex
Martins, Samantha M.
Holub, Oliver
Rocha, Surza L. G.
Valente, Richard H.
Ferreira, Ana G. C. Neves
Ferreira, Sérgio T.
Domont, Gilberto B.
Perales, Jonas
Affilliation: Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Toxinologia. Rio de Janeiro, RJ, Brasil.
Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica. Programa de Bioquímica e Biofísica Celular. Rio de Janeiro, RJ, Brasil.
Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica. Programa de Bioquímica e Biofísica Celular. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Toxinologia. Rio de Janeiro, RJ, Brasil / Universidade Federal do Rio de Janeiro. Instituto de Química. Departamento de Bioquímica. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Toxinologia. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Toxinologia. Rio de Janeiro, RJ, Brasil.
Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica. Programa de Bioquímica e Biofísica Celular. Rio de Janeiro, RJ, Brasil.
Universidade Federal do Rio de Janeiro. Instituto de Química. Departamento de Bioquímica. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Toxinologia. Rio de Janeiro, RJ, Brasil.
Abstract: We have investigated the folding of DM43, a homodimeric metalloproteinase inhibitor isolated from the serum of the South American opossum Didelphis marsupialis. Denaturation of the protein induced by GdnHCl (guanidine hydrochloride) was monitored by extrinsic and intrinsic fluorescence spectroscopy. While the equilibrium (un)folding of DM43 followed by tryptophan fluorescence was well described by a cooperative two-state transition, bis-ANS (4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonic acid) fluorescence measurements revealed an intensity maximum at the midpoint of the unfolding transition (2 M GdnHCl), indicating a partially folded intermediate state. We further investigated the DM43 intermediate stabilized at 2 M GdnHCl using size exclusion chromatography. This analysis revealed that the folding intermediate can be best described as partially folded DM43 monomers. Thermodynamic analysis of the GdnHCl-induced denaturation of DM43 revealed Gibbs free-energy changes of 13.57 kcal/mol for dimer dissociation and 1.86 kcal/mol for monomer unfolding, pointing to a critical role of dimerization as a determinant of the structure and stability of this protein. In addition, by using hydrostatic pressure (up to 3.5 kbar) we were able to stabilize partially folded states different from those stabilized in the presence of GdnHCl. Taken together, these results indicate that the conformational plasticity of DM43 could provide this protein with the ability to adapt its conformation to a variety of different environments and biological partners during its biological lifetime.
Keywords: Snake venom inhibitor
Protein folding
Intermediate
Conformational plasticity
keywords: Inibidor de veneno de cobra
Dobramento de proteínas
Intermediário
Plasticidade Conformacional
Issue Date: 2009
Publisher: Elsevier
Citation: CHAPEAUROUGE, Alex; et al. Conformational plasticity of DM43, a metalloproteinase inhibitor from Didelphis marsupialis: Chemical and pressure-induced equilibrium (un)folding studies. Biochimica et Biophysica Acta, v.1794, p.1379–1386, Mar. 2009.
DOI: 10.1016/j.bbapap.2009.03.010
ISSN: 1570-9639
Copyright: restricted access
Appears in Collections:IOC - Artigos de Periódicos

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