Molecular cloning and characterization of bile salt hydrolase from Lactobacillus fermentum

The gastrointestinal normal flora plays a vital role in human physiology, as it controls the host's nutritious, physiological, and immunological activities. In the human intestine, bile salt hydrolase (BSH), produced mainly by intestinal bacteria, catalyzes the hydrolysis of taurine or bile acid bound to glycine binding to amino acid and deconjugated bile acid. Deconjugated bile acids play an important role in the developing certain gastrointestinal diseases, such as gallstone formation, cholestasis, and colon cancer as well as blood cholesterol levels. However, the LAB are a group of microorganisms that mainly essensial for fermentative of dietary food in the market today. Because of the correlation with both BSH and human health, the structural and biochemical properties of BSHs require considerable research in order to understand their role in the regulation bacterial and host metabolism. The catalytic activities and substrate specificities of BSHs vary more. Some amino acids reported to be responsible for catalytic activity were also entirely kept intact across all BSH members of the same family. As an outcome, knowing and ability to understand the mechanism of action of the BSH enzymes of the probiotic microorganisms used in those treatments and fermentation is very vital. Despite the fact that Lactobacillus fermentum is likely one of the most beneficial probiotic bacteria strain on the market in the world today, little information is known about the mechanism of action and substrate preferences of this microorganism's BSH enzyme. In this study, the bsh gene encoding 326-amino acids from L. fermentum strain was cloned, expressed and characterized in Escherichia coli BLR(DE3) strain. An E. coli BLR(DE3)-pET22b expression system was used to produce rcombinant BSH. The recombinant BSH activity of L. fermentum was not determined qualitatively using the Direct Plate Assay with two different bile salts, glydeoxucocholic acid and taurodeoxycholic acid. In addition to this, 37 kDa BSH protein was not visible on SDS-PAGE analysis. Multiple sequence alignment revealed that BSH enzyme of L. fermentum had a one mutation (one codon deletion) on bsh gene. This mutation can affect the stability of BSH enzyme of L. fermentum. In this respect, L. fermentum cannot be a potential probiotic candidate since the capability of BSH activity is one of the criteria for the selection of the probiotics.