Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) genome sequence analysis revealed the presence of two genes that encode histone-like HU proteins (hlbA and hlbB) showing extensive similarity to other bacterial homologues. These genes were found to be extremely conserved among several L. bulgaricus strains. The hlbA gene was shown to be constitutively transcribed from a unique promoter (phlbA) during normal growth, whereas hlbB did not seem to be expressed under usual laboratory conditions. Using a reporter cassette in which the staphylococcal nuclease was fused at its N-terminus to the lactococcal signal peptide Usp45 (SP Usp45), we have demonstrated that phlbA promotes high expression of the reporter in L. bulgaricus, which correlated with an abundant secretion of the mature nuclease in the supernatant fraction. Quantification of the exported enzyme reveals a secretion level approximately threefold higher when the expression of the reporter was under the control of phlbA compared with the lactococcal usp45 promoter. Together, these results indicate that phlbA is suitable for gene expression in L. bulgaricus, that SP Usp45 is functionally recognized and processed by the L. bulgaricus secretion machinery and that the nuclease reporter gene can be used for the identification of exported products in this bacterium.
L-Arabinose isomerase stability is a crucial criterion for the industrial application of this biocatalyst. Noria and NoriaPG are capable of increasing the L-arabinose isomerase stability not only at high temperatures but also at low pH. Such results highlight, for the first time, the use of the Noria series of molecules for protein stabilization and activation.
Identification of stress proteins in Lactobacillus delbrueckii bulgaricus by two-dimensional electrophoresis.We studied Lactobacillus bulgaricus adaptation to an acid stress.In L. bulgaricus, adapted cells (30 min incubation at pH 4.75) were ~250 fold more tolerant to lethal acid stress (30 min at pH 3.6) than non adapted ones.In order to identify proteins induced during the adaptation, de novo protein synthesis was monitored by 35 S methionine labeling of exponentially growing cultures, under standard (pH 6) and adaptive (pH 4.75) conditions.After 2 dimensional (2-D) electrophoresis separation, the protein patterns were compared.At least 30 protein spots showed an increased radioactivity level under acidic conditions.We determined the N-terminal amino acid sequence of 3 highly induced proteins.By comparison to databases, they were identified as the heat shock proteins GroES, GroEL and DnaK.A differential induction was observed for GroES and GroEL, suggesting a post-transcriptionnal regulation mechanism.In this report we established a method for the study of acid stress response in L. bulgaricus.This method will also be useful for further studies on the adaptation of this bacterium to environmental variations. Lactobacillus bulgaricus / two dimensional electrophoresis / acid stress / adaptation / heat shock proteinRésumé -Nous avons étudié l'adaptation de la bactérie lactique Lactobacillus bulgaricus au stress acide.Nous avons montré que les bactéries cultivées en milieu « neutre » (pH 6) présentaient une forte létalité après transfert dans un milieu acide (choc acide à pH 3,6).En revanche, les bactéries préalablement adaptées dans un milieu acide non létal (pH 4,75) devenaient ~250 fois plus tolérantes au choc acide.Après marquage à la méthionine 35 S et séparation par électrophorèse bidimensionnelle (2-D), les profils protéiques obtenus en condition d'adaptation (pH 4,75) et en condition standard (pH 6) sont comparés.On observe une augmentation significative (>2) de l'intensité d'environ 30 spots pendant l'adaptation.La détermination des séquences peptidiques N-terminales de 3 protéines fortement induites à pH 4,75 permet d'identifier les protéines de choc thermique GroES, GroEL et
The role of the gut microbiota in health and disease is well recognized and the microbiota dysbiosis observed in many chronic diseases became a new therapeutic target. The challenge is to get a better insight into the functionality of commensal bacteria and to use this knowledge to select live biotherapeutics as new preventive or therapeutic products. In this study, we set up a screening approach to evaluate the functional capacities of a set of 21 strains isolated from the gut microbiota of neonates and adults. For this purpose, we selected key biological processes involved in the microbiome-host symbiosis and known to impact the host physiology i.e., the production of short-chain fatty acids and the ability to strengthen an epithelial barrier (Caco-2), to induce the release of the anti-inflammatory IL-10 cytokine after co-culture with human immune cells (PBMC) or to increase GLP-1 production from STC-1 endocrine cell line. This strategy highlighted fifteen strains exhibiting beneficial activities among which seven strains combined several of them. Interestingly, this work revealed for the first time a high prevalence of potential health-promoting functions among intestinal commensal strains and identified several appealing novel candidates for the management of chronic diseases, notably obesity and inflammatory bowel diseases.
Lactobacillus delbrueckii ssp. lactis and ssp. bulgaricus are lactic acid producing bacteria that are largely used in dairy industries, notably in cheese-making and yogurt production. An earlier in-depth study of the first completely sequenced ssp. bulgaricus genome revealed the characteristics of a genome in an active phase of rapid evolution, in what appears to be an adaptation to the milk environment. Here we examine for the first time if the same conclusions apply to the ssp. lactis, and discuss intra- and inter-subspecies genomic diversity in the context of evolutionary adaptation. Both L. delbrueckii ssp. show the signs of reductive evolution through the elimination of superfluous genes, thereby limiting their carbohydrate metabolic capacities and amino acid biosynthesis potential. In the ssp. lactis this reductive evolution has gone less far than in the ssp. bulgaricus. Consequently, the ssp. lactis retained more extended carbohydrate metabolizing capabilities than the ssp. bulgaricus but, due to high intra-subspecies diversity, very few carbohydrate substrates, if any, allow a reliable distinction of the two ssp. We further show that one of the most important traits, lactose fermentation, of one of the economically most important dairy bacteria, L. delbruecki ssp. bulgaricus, relies on horizontally acquired rather than deep ancestral genes. In this sense this bacterium may thus be regarded as a natural GMO avant la lettre. The dairy lactic acid producing bacteria L. delbrueckii ssp. lactis and ssp. bulgaricus appear to represent different points on the same evolutionary track of adaptation to the milk environment through the loss of superfluous functions and the acquisition of functions that allow an optimized utilization of milk resources, where the ssp. bulgaricus has progressed further away from the common ancestor.
The Serine Protease Inhibitors (Serpins) have been a focus of research by biomedical industries due to their critical role in human health. The use of serpin in the treatment of many diseases was widely investigated through the identification of new genes encoding these proteins in all kingdoms of life. The characterization of these genes revealed that they encoded proteins having low sequence homologies. Future developments are focusing not only on the protease inhibition activity, but also on the other effects due to the interactions of serpins with other components such as hormone transport. Here we give a concise overview of the most recent patents that have been reported in this field of research.
Silver nanoparticles capped with nine different sulphonated calix[n]arenes were tested for their anti-bacterial effects against B. subtilis and E. coli at an apparent concentration of 100 nM in calix[n]arene. The results show the para-sulphonato-calix[n]arenes are active against Gram positive bacteria and the derivatives having sulphonate groups at both para and alkyl terminal positions are active against Gram negative bacteria. The calix[6]arene derivative with only O-alkyl sulphonate groups shows bactericidal activity.
Lactococcus lactis, one of the most commonly used dairy starters, is often subjected to oxidative stress in cheese manufacturing. A comparative proteomic analysis was performed to identify the molecular modifications responsible for the robustness of three spontaneous H(2)O(2)-resistant (SpOx) strains. In the parental strain, glyceraldehyde-3-phosphate deshydrogenase (GAPDH) activity is ensured by GapB and the second GAPDH GapA is not produced in standard growth conditions. We showed that GapA was overproduced in the highly resistant SpOx2 and SpOx3 mutants. Its overproduction in the MG1363 strain led to an increased H(2)O(2) resistance of exponential growing cells. Upon H(2)O(2) exposure, GapB was fully inactivated by oxidation in the parental strain. In SpOx mutants, it partly remained in the reduced form sustaining partially GAPDH activity. The analysis of gapA disruption in these SpOx strains indicated that additional unraveled mechanisms likely contribute to the resistance phenotype. In the SpOx1 mutant, the arginine deiminase pathway was found to be upregulated and disruption of arcA or arcB genes abolished H(2)O(2) resistance. We concluded that arginine consumption was directly responsible for the SpOx1 phenotype. Finally, these results suggest that sustaining energy supply is a major way of leading to oxidative stress resistance in L. lactis.
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