Structural insights on the efficient catalysis of hydroperoxide reduction by Ohr: Crystallographic and molecular dynamics approaches

Autor(es)

Piccirillo, Erika ^[1] ; Alegria, Thiago G. P. ^[2] ; Discola, Karen F. ^[2] ; Cussiol, Jose A. R. R. ^{[3, 2]} ; Domingos, Renato M. ^[2] ; de Oliveira, Marcos A. ^[4] ; de Rezende, Leandro ^[1] ; Netto, Luis E. S. ^[2] ; Amaral, Antonia T-do ^[1]

Afiliação do(s) autor(es)

^[1] Univ Sao Paulo, Inst Quim, Dept Quim Fundamental, Sao Paulo, SP - Brazil

^[2] Univ Sao Paulo, Inst Biociencias, Dept Genet & Biol Evolut, Sao Paulo, SP - Brazil

^[3] Univ Fed Sao Paulo, Dept Bioquim, Sao Paulo, SP - Brazil

^[4] Univ Estadual Paulista, Inst Biociencias, Campus Litoral Paulista, Sao Vicente, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: PLoS One; v. 13, n. 5 MAY 21 2018.
• Citações Web of Science: 1

Resumo

Organic hydroperoxide resistance (Ohr) enzymes are highly efficient Cys-based peroxidases that play central roles in bacterial response to fatty acid hydroperoxides and peroxynitrite, two oxidants that are generated during host-pathogen interactions. In the active site of Ohr proteins, the conserved Arg (Arg19 in Ohr from Xylella fastidiosa) and Glu (Glu51 in Ohr from Xylella fastidiosa) residues, among other factors, are involved in the extremely high reactivity of the peroxidatic Cys (C-p) toward hydroperoxides. In the closed state, the thiolate of C-p is in close proximity to the guanidinium group of Arg19. Ohr enzymes can also assume an open state, where the loop containing the catalytic Arg is far away from C-p and Glu51. Here, we aimed to gain insights into the putative structural switches of the Ohr catalytic cycle. First, we describe the crystal structure of Ohr from Xylella fastidiosa (XfOhr) in the open state that, together with the previously described XfOhr structure in the closed state, may represent two snapshots along the coordinate of the enzyme-catalyzed reaction. These two structures were used for the experimental validation of molecular dynamics (MD) simulations. MD simulations employing distinct protonation states and in silico mutagenesis indicated that the polar interactions of Arg19 with Glu51 and C-p contributed to the stabilization of XfOhr in the closed state. Indeed, C-p oxidation to the disulfide state facilitated the switching of the Arg19 loop from the closed to the open state. In addition to the Arg19 loop, other portions of XfOhr displayed high mobility, such as a loop rich in Gly residues. In summary, we obtained a high correlation between crystallographic data, MD simulations and biochemical/enzymatic assays. The dynamics of the Ohr enzymes are unique among the Cys-based peroxidases, in which the active site Arg undergoes structural switches throughout the catalytic cycle, while C-p remains relatively static. (AU)

• Processo FAPESP: 14/01614-5 - Aplicação de dinâmica molecular para melhorar modelos farmacofóricos da protease NS2B/NS3 do DENV, considerando a flexibilidade conformacional da protease, e para validar modelos de homologia para diferentes serotipos de proteases
• Beneficiário: Erika Piccirillo
• Linha de fomento: Bolsas no Exterior - Estágio de Pesquisa - Doutorado Direto
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• Linha de fomento: Bolsas no Brasil - Mestrado
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• Linha de fomento: Bolsas no Brasil - Doutorado Direto
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