RESUMO
Trace amounts of the carcinogenic ethyl carbamate can appear in wine as a result of a reaction between ethanol and citrulline, which is produced from arginine degradation by some bacteria used in winemaking. In this study, arginine deiminase (ADI) pathway genes were evaluated in 44 Oenococcus oeni strains from wines originating from several locations in order to establish the relationship between the ability of a strain to degrade arginine and the presence of related genes. To detect the presence of arc genes of the ADI pathway in O. oeni, pairs of primers were designed to amplify arcA, arcB, arcC and arcD1 sequences. All strains contained these four genes. The same primers were used to confirm the organization of these genes in an arcABCD1 operon. Nevertheless, considerable variability in the ability to degrade arginine among these O. oeni strains was observed. Therefore, despite the presence of the arc genes in all strains, the expression patterns of individual genes must be strain dependent and influenced by the different wine conditions. Additionally, the presence of arc genes was also determined in the 57 sequenced strains of O. oeni available in GenBank, and the complete operon was found in 83% of strains derived from wine. The other strains were found to lack the arcB, arcC and arcD genes, but all contained sequences homologous to arcA, and some of them had also ADI activity.
Assuntos
Arginina/metabolismo , Hidrolases/metabolismo , Oenococcus/enzimologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Hidrolases/genética , Oenococcus/classificação , Oenococcus/genética , Oenococcus/metabolismo , Óperon , Vinho/microbiologiaRESUMO
Trace amounts of the carcinogen ethyl carbamate can appear in wine by the reaction of ethanol with compounds such as citrulline and carbamyl phosphate, which are produced from arginine degradation by some wine lactic acid bacteria (LAB). In this work, the presence of arc genes for the arginine-deiminase pathway was studied in several strains of different species of LAB. Their ability to degrade arginine was also studied. To detect the presence of arc genes, degenerate primers were designed from the alignment of protein sequences in already sequenced LAB. The usefulness of these degenerate primers has been proven by sequencing some of the amplified PCR fragments and searching for homologies with published sequences of the same species and related ones. Correlation was found between the presence of genes and the ability to degrade arginine. Degrading strains included all heterofermentative lactobacilli, Oenococcus oeni , Pediococcus pentosaceus , and some strains of Leuconostoc mesenteroides and Lactobacillus plantarum .
Assuntos
Proteínas de Bactérias/genética , Ácido Láctico/metabolismo , Lactobacillaceae/genética , Uretana/metabolismo , Vinho/microbiologia , Proteínas de Bactérias/metabolismo , Lactobacillaceae/metabolismo , Dados de Sequência Molecular , Reação em Cadeia da PolimeraseRESUMO
The goal of this study was to analyze how the profiles of membrane proteins of Oenococcus oeni change under particular stress conditions of wine. Sodium dodecyl sulfate polyacrylamide gel electrophoresis protein profiles of membrane fraction showed that a 40-kDa protein was overexpressed in the presence of SO2. The sequence of its N-terminal fragment showed a significant identity with glyceraldehyde-3-phosphate dehydrogenases (GAPDHs), but the protein showed no GAPDH activity. This sequence was compared with those of other GAPDHs with ClustalW alignment, and it was found to be somewhat similar to that of the cell-wall and membrane proteins of other lactic acid bacteria.
Assuntos
Proteínas de Bactérias/análise , Gliceraldeído-3-Fosfato Desidrogenases/análise , Lactobacillaceae/enzimologia , Proteínas de Membrana/análise , Sequência de Aminoácidos , Proteínas de Bactérias/biossíntese , Gliceraldeído-3-Fosfato Desidrogenases/biossíntese , Lactobacillaceae/crescimento & desenvolvimento , Proteínas de Membrana/biossíntese , Dados de Sequência Molecular , Análise de Sequência de Proteína/métodos , Vinho/microbiologiaRESUMO
The effects of different yeast starters and SO(2) addition on malolactic fermentation in a new winery were evaluated by a molecular approach in three vintages. Alcoholic fermentations with 40 and 100mgl(-1) SO(2) were carried out, followed by uninoculated malolactic fermentations. Isolated colonies of Oenococcus oeni obtained from samples throughout the vinification were identified and typified by multiplex RAPD-PCR. This made it possible to monitor the population dynamics and follow the proportion of as many as 29 different indigenous strains. In one of the vintages, O. oeni strains were more inhibited when a specific yeast starter was used.
Assuntos
Cocos Gram-Positivos/classificação , Cocos Gram-Positivos/crescimento & desenvolvimento , Dióxido de Enxofre , Vinho/microbiologia , Leveduras , Impressões Digitais de DNA , DNA Bacteriano/análise , DNA Bacteriano/isolamento & purificação , Fermentação , Microbiologia de Alimentos , Cocos Gram-Positivos/isolamento & purificação , Cocos Gram-Positivos/metabolismo , Técnica de Amplificação ao Acaso de DNA PolimórficoRESUMO
Malolactic fermentation (MLF) is carried out by Oenococcus oeni under very harsh conditions. This paper shows that stress compounds in wine such as SO(2), fatty acids and copper have an inhibitory effect on cell growth and MLF duration, and relates this effect to an inhibition of ATPase activity. Of the stress compounds, SO(2) and dodecanoic acid had the strongest effect, decreasing the ATPase specific activity to 37% and 58%, respectively. It can be concluded that ATPase is a good indicator of the physiological state of the cells and their ability to lead MLF.
