Lipid aldehyde hydrophobicity affects apo-SOD1 modification and aggregation.

Unsaturated lipids are oxidized by reactive oxygen species and enzymes, leading to the increased formation of lipid hydroperoxides and several electrophilic products. Lipid-derived electrophiles can modify macromolecules, such as proteins, resulting in the loss of function and/or aggregation. The accumulation of Cu,Zn-superoxide dismutase (SOD1) aggregates has been associated with familial cases of amyotrophic lateral sclerosis (ALS). The protein aggregation mechanisms in motor neurons remain unclear, although recent studies have shown that lipids and oxidized lipid derivatives may play roles in this process. Here, we aimed to compare the effects of different lipid aldehydes on the induction of SOD1 modifications and aggregation, in vitro. Human recombinant apo-SOD1 was incubated with 4-hydroxy-2-hexenal (HHE), 4-hydroxy-2-nonenal (HNE), 2-hexen-1-al (HEX), 2,4-nonadienal (NON), 2,4-decadienal (DEC), or secosterol aldehydes (SECO-A or SECO-B). High-molecular-weight apo-SOD1 aggregates dramatically increased in the presence of highly hydrophobic aldehydes (LogPcalc > 3). Notably, several Lys residues were modified by exposure to all aldehydes. The observed modifications were primarily observed on Lys residues located near the dimer interface (K3 and K9) and at the electrostatic loop (K122, K128, and K136). Moreover, HHE and HNE induced extensive apo-SOD1 modifications, by forming Schiff bases or Michael adducts with Lys, His, and Cys residues. However, these aldehydes were unable to induce large protein aggregates. Overall, our data shed light on the importance of lipid aldehyde hydrophobicity on the induction of apo-SOD1 aggregation and identified preferential sites of lipid aldehyde-induced modifications.

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

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 (Cp) toward hydroperoxides. In the closed state, the thiolate of Cp 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 Cp 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 Cp contributed to the stabilization of XfOhr in the closed state. Indeed, Cp 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 Cp remains relatively static.

Produtividade de criadouro de Aedes albopictus em ambiente urbano / Productivity of container-breeding Aedes albopictus in an urban environment

O encontro de Aedes albopictus na cidade de Cananéia, regiäo Sudeste do Estado de Säo Paulo, Brasil, ensejou a ocasiäo de realizar observaçöes que visassem a avaliar a produtividade de criadouro grande e permanente. Objetiva, após selecionar o habitat a ser estudado, avaliar a contribuiçäo para a densidade local do mosquito. Em área predeterminada procedeu-se a levantamento de criadouros potenciais. Constatada a presença da espécie, foi selecionado um dos recipientes que preenchia os requisitos desejados. O acompanhamento foi feito de maneira ininterrupta, no período de novembro de 1996 a maio de 1997. As observaçöes obedeceram a ritmo quinzenal retirando, cada vez, amostra da água correspondente a 0,14, ou seja, um sétimo do volume total de 70 litros. Procurou-se coletar, identificar e numerar, por sexo, as pupas existentes. Concomitantemente, procedeu-se à captura de formas adultas. Foi utilizada a isca humana das 15:00 às 18:00h, instalada a cerca de 6 metros do mencionado criadouro. Finda essa coleta, foi feita aspiraçäo com 30 min. de duraçäo em locais de abrigo representados pela abundante vegetaçäo circunjacente. Nas coletas de formas imaturas do criadouro, o Ae. albopictus compareceu com 44,9 por cento. Ao longo de 15 amostras regularmente realizadas obteve-se a média de 31,13 pupas pertencentes a essa espécie. O índice de emergência(E) foi de 2,1.A multiplicaçäo desse valor por sete forneceu a média diária de 14,7 fêmeas. Nas coletas de adultos desse sexo, a média de Williams para a isca humana foi de 30,7, enquanto a densidade média horária da aspiraçäo dos locais de abrigo foi de 9,2. O cálculo do acúmulo diário concluiu pela presença de 22,8 fêmeas, por dia, capazes de freqüentar a isca humana, nessa situaçäo e condiçöes. A contagem de pupas possibilitou estimar a produtividade de criadouro de Ae. albopictus, tipo grande (dez litros ou mais) e de caráter permanente. A água acumulada no recipiente estudado apresentou-se rica em matéria orgânica, predominantemente de natureza vegetal. Näo há como compará-lo a reservatórios destinados ao armazenamento de água para uso doméstico. A falta de manutençäo desses reservatórios poderá contribuir para levar a situaçäo, se näo idêntica, pelo menos próxima da evidenciada no trabalho. Embora se trate de espécie até agora näo incriminada como vetora, é de se admitir que as observaçöes encontradas possam ser utilizadas no programa de erradicaçäo de Ae. albopictus que está em em curso no Brasil