Comparative Proteomic Analysis of Lactobacillus plantarum WCFS1 and ΔctsR Mutant Strains Under Physiological and Heat Stress Conditions (original) (raw)
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BMC Microbiology, 2011
Background Lactic acid bacteria are commonly marketed as probiotics based on their putative or proven health-promoting effects. These effects are known to be strain specific but the underlying molecular mechanisms remain poorly understood. Therefore, unravelling the determinants behind probiotic features is of particular interest since it would help select strains that stand the best chance of success in clinical trials. Bile tolerance is one of the most crucial properties as it determines the ability of bacteria to survive in the small intestine, and consequently their capacity to play their functional role as probiotics. In this context, the objective of this study was to investigate the natural protein diversity within the Lactobacillus plantarum species with relation to bile tolerance, using comparative proteomics. Results Bile tolerance properties of nine L. plantarum strains were studied in vitro. Three of them presenting different bile tolerance levels were selected for comparative proteomic analysis: L. plantarum 299 V (resistant), L. plantarum LC 804 (intermediate) and L. plantarum LC 56 (sensitive). Qualitative and quantitative differences in proteomes were analyzed using two-dimensional electrophoresis (2-DE), tryptic digestion, liquid chromatography-mass spectrometry analysis and database search for protein identification. Among the proteins correlated with differences in the 2-DE patterns of the bacterial strains, 15 have previously been reported to be involved in bile tolerance processes. The effect of a bile exposure on these patterns was investigated, which led to the identification of six proteins that may be key in the bile salt response and adaptation in L. plantarum: two glutathione reductases involved in protection against oxidative injury caused by bile salts, a cyclopropane-fatty-acyl-phospholipid synthase implicated in maintenance of cell envelope integrity, a bile salt hydrolase, an ABC transporter and a F0F1-ATP synthase which participate in the active removal of bile-related stress factors. Conclusions These results showed that comparative proteomic analysis can help understand the differential bacterial properties of lactobacilli. In the field of probiotic studies, characteristic proteomic profiles can be identified for individual properties that may serve as bacterial biomarkers for the preliminary selection of strains with the best probiotic potential.
Molecular & Cellular Proteomics, 2011
Lactobacillus rhamnosus GG (GG) is a widely used and intensively studied probiotic bacterium. Although the health benefits of strain GG are well documented, the systematic exploration of mechanisms by which this strain exerts probiotic effects in the host has only recently been initiated. The ability to survive the harsh conditions of the gastrointestinal tract, including gastric juice containing bile salts, is one of the vital characteristics that enables a probiotic bacterium to transiently colonize the host. Here we used gene expression profiling at the transcriptome and proteome levels to investigate the cellular response of strain GG toward bile under defined bioreactor conditions. The analyses revealed that in response to growth of strain GG in the presence of 0.2% ox gall the transcript levels of 316 genes changed significantly (p < 0.01, t test), and 42 proteins, including both intracellular and surface-exposed proteins (i.e. surfome), were differentially abundant (p < 0.01, t test in total proteome analysis; p < 0.05, t test in surfome analysis). Protein abundance changes correlated with transcriptome level changes for 14 of these proteins. The identified proteins suggest diverse and specific changes in general stress responses as well as in cell envelope-related functions, including in pathways affecting fatty acid composition, cell surface charge, and thickness of the exopolysaccharide layer. These changes are likely to strengthen the cell envelope against bile-induced stress and signal the GG cells of gut entrance. Notably, the surfome analyses demonstrated significant reduction in the abundance of a protein catalyzing the synthesis of exopolysaccharides, whereas a protein dedicated for active removal of bile compounds from the cells was up-regulated. These findings suggest a role for these proteins in facilitating the well founded interaction of strain GG with the host mucus in the presence of sublethal doses of bile. The significance of these findings in terms of the functionality of a probiotic bacterium is discussed. Molecular & Cellular
Lactobacillus fermentum NCDC 400 is a commonly used and a comprehensively studied probiotic organism. The distinctive capability to endure the harsh environment of the gastrointestinal tract owing to the presence of bile salts in gastric juice facilitates gut microbiota, especially the probiotic organisms to transiently inhabit the host gut. In the present study, Label-Free Quantification (LFQ) approach has been employed to analyze the expression pattern of Lactobacillus fermentum NCDC 400 strain proteins, under bile acid stress, using high-resolution mass spectrometer connected to nano-liquid chromatography (LC) system. We report the identification of a total of 538 differentially expressed (DE) proteins in response to 1.2% bile salt which is required for the growth of this bacterium. Among the DE proteins, 80 were found to be up-regulated, with greater than 1.3 fold change vis-à-vis 107 proteins which were down-regulated with < 0.76 fold change (p < 0.05). The functions of down-regulated proteins were largely unknown nevertheless; the putative functions of the up-regulated proteins were categorized into categories viz. stress response, DNA repair, peptidoglycan biosynthesis, amino acids metabolism, signal transduction, transcription, translation, and carbohydrate metabolism. These results suggest that the differentially expressed proteins provide the tolerance towards the various gastrointestinal challenges and involved in bile acid stress response mechanism.
Applied and Environmental Microbiology, 2007
Bile salts are natural detergents that facilitate the digestion and absorption of the hydrophobic components of the diet. However, their amphiphilic nature makes them very inhibitory for bacteria and strongly influences bacterial survival in the gastrointestinal tract. Adaptation to and tolerance of bile stress is therefore crucial for the persistence of bacteria in the human colonic niche. Bifidobacterium animalis subsp. lactis, a probiotic bacterium with documented health benefits, is applied largely in fermented dairy products. In this study, the effect of bile salts on proteomes of B. animalis subsp. lactis IPLA 4549 and its bile-resistant derivative B. animalis subsp. lactis 4549dOx was analyzed, leading to the identification of proteins which may represent the targets of bile salt response and adaptation in B. animalis subsp. lactis. The comparison of the wild-type and the bile-resistant strain responses allowed us to hypothesize about the resistance mechanisms acquired by the derivative resistant strain and about the bile salt response in B. animalis subsp. lactis. In addition, significant differences in the levels of metabolic end products of the bifid shunt and in the redox status of the cells were also detected, which correlate with some differences observed between the proteomes. These results indicate that adaptation and response to bile in B. animalis subsp. lactis involve several physiological mechanisms that are jointly dedicated to reduce the deleterious impact of bile on the cell's physiology.
Proteomic and transcriptomic analysis of the response to bile stress of L. casei BL23
Microbiology (Reading, England), 2012
Lactobacillus casei is a lactic acid bacterium commonly found in the gastrointestinal tract of animals and some strains are used as probiotics. The ability of probiotic strains to survive the passage through the gastrointestinal tract is considered a key factor for their probiotic action. Therefore, tolerance to bile salts is a desirable feature for probiotic strains. In this study we have characterized the response of L. casei BL23 to bile by a transcriptomic and proteomic approach. The analysis revealed that exposition to bile induced changes in the abundance of 52 proteins and the transcript levels of 67 genes. The observed changes affected genes and proteins involved the stress response, fatty acid and cell wall biosynthesis, metabolism of carbohydrates, transport of peptides, coenzyme levels, membrane H+-ATPase, and a number of uncharacterized genes and proteins. These data provide new insight into the mechanisms that enable L. casei BL23 to cope with bile stress.
Exploring the Bile Stress Response of Lactobacillus mucosae LM1 through Exoproteome Analysis
Molecules
Lactobacillus sp. have long been studied for their great potential in probiotic applications. Recently, proteomics analysis has become a useful tool for studies on potential lactobacilli probiotics. Specifically, proteomics has helped determine and describe the physiological changes that lactic acid bacteria undergo in specific conditions, especially in the host gut. In particular, the extracellular proteome, or exoproteome, of lactobacilli contains proteins specific to host– or environment–microbe interactions. Using gel-free, label-free ultra-high performance liquid chromatography tandem mass spectrometry, we explored the exoproteome of the probiotic candidate Lactobacillus mucosae LM1 subjected to bile treatment, to determine the proteins it may use against bile stress in the gut. Bile stress increased the size of the LM1 exoproteome, secreting ribosomal proteins (50S ribosomal protein L27 and L16) and metabolic proteins (lactate dehydrogenase, phosphoglycerate kinase, glyceralde...
Beneficial bile acid metabolism from Lactobacillus plantarum of food origin
Scientific Reports
Bile acid (BA) signatures are altered in many disease states. BA metabolism is an important microbial function to assist gut colonization and persistence, as well as microbial survival during gastro intestinal (GI) transit and it is an important criteria for potential probiotic bacteria. Microbes that express bile salt hydrolase (BSH), gateway BA modifying enzymes, are considered to have an advantage in the gut. This property is reported as selectively limited to gut-associated microbes. Food-associated microbes have the potential to confer health benefits to the human consumer. Here, we report that food associated Lactobacillus plantarum strains are capable of BA metabolism, they can withstand BA associated stress and propagate, a recognised important characteristic for GIT survival. Furthermore, we report that these food associated Lactobacillus plantarum strains have the selective ability to alter BA signatures in favour of receptor activation that would be beneficial to humans. ...
Exoproteome Perspective on the Bile Stress Response of Lactobacillus johnsonii
Proteomes
Probiotics must not only exert a health-promoting effect but also be capable of adapting to the harsh environment of the gastrointestinal (GI) tract. Probiotics in the GI tract must survive the cell wall-disrupting effect of bile acids. We investigated the exoproteome of Lactobacillus johnsonii PF01 and C1-10 under bile stress. A comparative analysis revealed the similarities between the two L. johnsonii exoproteomes, as well as their different responses to bile. The large number of metabolic proteins in L. johnsonii revealed its metabolic adaptation to meet protein synthesis requirements under bile stress. In addition, cell wall modifications occurred in response to bile. Furthermore, some extracellular proteins of L. johnsonii may have moonlighting function in the presence of bile. Enolase, L-lactate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, triosephosphate isomerase, 50s ribosomal protein L7/L12, and cellobiose-specific phosphotransferase system (PTS) sugar transpo...
Probiotic Properties and Bile Salt Hydrolase Activity of Some Isolated Lactic Acid Bacteria
Egyptian Journal of Microbiology, 2017
E IGHT lactic acid bacterial (LAB) isolates were obtained from food and non-food sources and identified by 16S rRNA gene sequence analysis. Based on the sequencing results, the isolates belong to two species of lactobacilli, Lactobacillus plantarum and L. rhamnosus. These strains were then compared with a reference strain, Lactobacillus casei, to assess various probiotic properties, such as haemolytic activity, histamine formation ability, stress tolerance under certain stress conditions, antibiotic susceptibility and in vitro adhesion ability. Moreover, bile salt hydrolase (BSH) activity was evaluated both qualitatively and quantitatively. The results showed that none of the isolates demonstrated any haemolytic activity or histamine formation. The isolated strains were also tolerant to acidic and alkaline conditions (pH 2.5, 3.5 and 9) for 3 and 6 h, as well as osmotic (3 M NaCl) and heat (55 and 70°C) stress, but were more responsive to oxidative and bile stress. The bacterial isolates also expressed high amounts of BSH, ranging from 90 to 142 U/mg in active cells, compared to L. casei (74 U/mg), which may be useful in cholesterol reduction. All bacterial isolates were resistant to vancomycin and susceptible to amoxicillin, cloxacillin and penicillin. All isolates were also highly hydrophobic (>70%), indicating that they are not easily flushed from the intestines.