Preliminary Study On The Biological Functions Of Adhesion Protein From Bifidobacterium

Bifidobacterium spp., the very important physiological bacterium in theintestinal tract of human and animals, play a key role in digestion, nutritionmetabolism and immunoregulation of the host. Adhesion and engraftment in thedigestive tract were the premise. This study aims to make a preliminary study on thebiological functions of adhesion protein from Bifidobacterium, and provide basicresearch materials for the exploration of the mechanism of cross-reaction betweenprobiotics and host.In this study, adhesion protein PP2and PP4were expressed in E. coli DH5αpGEX-4T-PP2and E. coli DH5α pGEX-4T-PP4by induction with IPTG, and thenpurified with GST affinity column. We verified the adhesion function of twoadhesion proteins PP2and PP4visually via the fluorescence microscope afterlabelled with quantum dot, and evaluated their adhesion effect on probiotics byplate counting method. The changing of mRNA expression in intestinal epithelialcells led by two adhesion proteins was evaluated by real time quantitative PCR. Theresults showed that the key factors related to immune regulation (such as JNK, p38,IL1R etc), cytoskeleton structure (such as GAB2, PAK etc), cell proliferationandtumor formation (such as Erbin, TRAIL, TGFβ etc) changed in different levels. Therelationship between the expression quantity changes of factors in signalingpathways and their related specific function is so complex, needs to be analyzed andexplored further.This study verified the inhibition effect of adhesion protein PP2, PP4againstthe adherence of Escherichia coli O157: H7, Salmonella typhimuriumon andListeria monocytogenes to epicelial cells by gram staining and plate countingmethods. The effects of adhesion inhibition were positively correlated with theadhesion proteins concentrations. The adhesion inhibition against Listeriamonocytogenes was the most pronounced (p<0.01), and adhesion proteins canobviously reduce its degree of antibiotic resistance. The mRNA expression quantity changes of Listeria monocytogenes after incubated with adhesion protein PP2andPP4were both evaluated by real time quantitative PCR. RPS7, RPL16expressionquantity reduced and the expression quantity of ASP, MUO, RPL29and GPSPincreased in different levels, and GPSP was related with the PBP synthesis.This study also screened some suspected adhesion proteins of Bifidobacteriumbifidum, Bifidobacterium infantis and Bifidobacterium adolescentis by the magneticbeads method in order to provide sufficient materials for further study.The research model established in this study can effectively guide the functionstudy of other adhesion proteins and the screening work of adhesive bacterium, itcan also provide more reference for the further study on the biological functions ofadhesive factors and the interaction mechanism of probiotics with the host.