engleski

Functional characterisation of the S-layer surface protein of Lactobacillus brevis D6

Over 200 lactic acid bacteria (LAB) strains were isolated from autochthonous fresh cheeses and deposited in the Collection of Lactic Acid Bacteria (ZBMK) of Laboratory of Antibiotic, Enzyme, Probiotic and Starter Cultures Technologies, Faculty of Food Technology and Biotechnology, in order to selected Lactobacillus strains with important functional properties. After phenotypic characterisation, selected strains were assayed for their autoaggregation capacities and S-layer protein presence on bacterial cell surface by SDS-PAGE analysis, as these proteins are appointed to play important biological roles in Lactobacillus strains. Further research revealed that the autochthonous strain D6, which is identified as Lactobacillus brevis, is an S-layer protein carrying strain. This strain displayed the highest percentage of autoaggregation (70.37% ± 5.79 after 5h) among tested LAB strains, which has been affected after 5M guanidine hydrochloride removal of S-layer proteins (54.87±7.13 after 5h). Since S-layer proteins have been identified, between LAB, only among representatives of Lactobacillus species, it was of the special interest to study its functional role in Lb. brevis D6. Subsequently, the capacities of the Lb. brevis D6 to adhere to extracellular matrix proteins (ECM) collagen, fibronectin and lamin and to human intestinal epithelial Caco-2 cell lines in vitro were assessed. Removal of the intact S-layer proteins from the bacterial cell surface, reduced the adhesion of Lb. brevis D6 to fibronectin, collagen and laminin. Binding tests showed that in vitro adhesion to ECM proteins of Lb. brevis D6, is of a proteinase K-sensitive manner. Additionally, coaggregation abilities, pathogen exclusion and survival in simulated conditions of gastrointestinal tract (GIT) and during freeze drying of Lb. brevis D6, before and after guanidine hydrochloride extraction of S-layer proteins from cell surface, were examined. The results suggest that S-layer protein enhances certain probiotic activities, especially adhesion to different surfaces of Lb. brevis D6.

adhesion; extracellular matrix proteins; Lactobacillus brevis; probiotic; S-layer proteins

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