RESUMO
The molecular dynamics of the Pseudomonas fluorescens 07A metalloprotease in the presence of structural Ca2+ and Mn2+ ions was evaluated. Seven Ca2+ ions are primarily bound to the C-terminus, while a divalent cation is located at the catalytic site, acting as a cofactor. The observed enzyme's experimental activity suggests that Mn2+ could compete for the active site of the enzyme with Ca2+, Zn2+ or other divalent cations, thus providing greater catalytic power to the enzyme. Our molecular dynamics simulations suggest that these ions partially protect the enzyme's structure from thermal denaturation. Moreover, our simulations have shown a collective movement of opening-closing of the active-site in simulations with structural Ca2+ and Mn2+ ions bound, leading to a proposal of a dynamical model of P. fluorescens 07A metalloprotease active and inactive conformations. These findings can support the development of measures to control the activity of P. fluorescens and other spoilage microorganism proteases.
Assuntos
Metaloproteases/metabolismo , Pseudomonas fluorescens/enzimologia , Sítios de Ligação , Cálcio/química , Cálcio/metabolismo , Domínio Catalítico , Cátions Bivalentes/química , Metaloproteases/química , Simulação de Dinâmica Molecular , Análise de Componente Principal , Zinco/química , Zinco/metabolismoRESUMO
Porcine circovirus-2 (PCV2) is the etiologic agent of several diseases in pigs, including multi-systemic wasting syndrome (PMWS). In this work, a new mutant PCV2b was isolated from PMWS-affected pigs on a Brazilian farm. Its genome showed high sequence similarity (>99% identity) to those from a group of emerging mutants isolated from cases of PMWS outbreaks in vaccinated pigs in China, the USA and South Korea. Here, we show that these isolates share a combination of low-frequency substitutions (single amino acid polymorphisms with a frequency of ≤25%) in the viral capsid protein, mainly in regions of immunoprotective epitopes, and an additional lysine residue at position 234. These isolates were phylogenetically grouped in the PCV2b clade, reinforcing the idea of the emergence of a new group of mutants PCV2b associated with outbreaks worldwide. The identification of these polymorphisms in the viral capsid highlights the importance of considering these isolates for the development of more-effective vaccines.
Assuntos
Substituição de Aminoácidos , Proteínas do Capsídeo/genética , Infecções por Circoviridae/veterinária , Circovirus/genética , Epitopos/genética , Síndrome Definhante Multissistêmico de Suínos Desmamados/virologia , Sequência de Aminoácidos , Animais , Brasil , Proteínas do Capsídeo/química , Proteínas do Capsídeo/imunologia , Infecções por Circoviridae/virologia , Circovirus/classificação , Circovirus/imunologia , Circovirus/isolamento & purificação , Epitopos/química , Epitopos/imunologia , Dados de Sequência Molecular , Filogenia , Polimorfismo de Nucleotídeo Único , SuínosRESUMO
Three porcine circovirus-2 strains were isolated from pigs on a Brazilian farm during an outbreak, indicating a vaccine failure. They present identical genomic sequences, with high identities to other isolates that were also related to vaccination failures, supporting the recent theory about an antigen drift being associated with vaccine failures throughout the world.