Adhesion And Protection Intestinal Epithelial Cells Of Lactobacillus Surface Layer Protein

Surface layer protein is one of major components that lactobacillus exerttheir probiotic. It gets more and more attention due to variety amounts andmultifunction. Investigations suggest that surface layer protein is the majorfactor that lactobacillus exert adhesion function which has decisive action ofintestinal protection. In recent years there is lack of study on purified surfacelayer protein and its protection mechanism of intestinal cells. Protective effect tointestinal epithelial cells was studied with purified surface layer protein in thisissue.In this issue, expression quantity of surface layer protein of Lactobacillusparacasei subsp. paracasei M5-L and Lactobacillus casei Q8-L were studiedunder different culture conditions. Both purified surface layer proteins by saltfractionation have single band at45kDa manifested by SDS-PAGE. Majorprotein from M5-L belongs to surface antigen and from Q8-L belongs toelongation factor manifested by LC/Q-TOF MS/MS, extract proteins from M5-Land Q8-L are surface layer protein proved by the results above.HT-29cells were labeled by Sulfo-NHS-Biotin (l.0mg/mL)and hybridizewith Surface layer proteins separated by electrophoresis.Surface layer proteins,from M5-L and Q8-L can specific binding to receptors on cytomembrane, haveadhesiveness. M5-L and Q8-L have smoothness and equality surface under thecondition that remain surface layer protein, whereas have relative rough andasymmetry surface observed by atomic force microscope; adhesive force andrelative elastic force of M5-L and Q8-L are1.41nN and0.48nN,18.82and27.94under the condition that remain surface layer protein, whereas are0.70nNand1.28nN,27.31and9.54respectively. Inhibition effect of surface layerprotein on S. sonnei to HT-29cells is measured by exclude, compete and replaceexperiments, the results showed that at the concentration of25?g/mL and400?g/mL of surface layer proteins from M5-L and Q8-L have the best effects,inhibition ratio between20%and50%.According to Hoechst33258HT-29cells were dyed and observed thencalculate the apoptosis ratio of cells. Apoptosis ratio of only add S. sonnei,surface layer protein from M5-L, Q8-L, S. sonnei with M5-L and S. sonnei withQ8-L are100.00%,5.00%,10.67%,37.67%and47.67%respectively. Changesof expression sites and distributions of tight junction ZO-1and Occludin manifested that surface layer protein from M5-L and Q8-L has inhibitory effectto S. sonnei on damage the ZO-1and Occludin of intestinal epithelial cellsobserved by immunofluorescence.