Molecular determinants of bacterial probiotic effects: the case of Bifidobacterium bifidum

Introduction Several beneficial effects on human health have been attributed to either commensal or food-derived probiotic bacteria, and their role in the modulation of the hostimmune response has been widely demonstrated. This research field is at the moment the most promising as for addressing the interplay between probiotics andthe host organisms.The identification of a number of molecular components that affect the host immune system activities may represent a powerful tool for developing new approachesboth in the field of immunologic diseases/deficiencies and in the treatment of colorectal cancer. Previous works of some of us demonstrate the immunomodulatory attitude of Bifidobacterium bifidum MIMBb75. In particular B. bifidum can render dendritic cells able to activate natural killer cells. Preliminary also showed that the immunomodulatory properties identified in the strain mentioned above might be associated with surface-exposed bacterial proteins. Therefore, identification and characterization of surface-exposed proteins in these bacterial strains seems relevant as for predicting their ability to interact with the host and to modulate immune system responses. Results In this work we proved that that the Cysteine Histidine-dependent Amidohydrolase/Peptidase (CHAP) domain of TgaA, a membrane associate protein from Bifidobacterium bifidum MIMBb75, can induce the production and release of interleukin (IL)-2 and Tumor Necrosis Factor (TNF)-α from mice dendritic cells (DC). In particular, we purified the recombinant protein TgaA and its two domains (LT and CHAP) from E. coli by means of NiNTA affinity columns, followed by RP-HPLC steps that proved essential for removing residual LPS and extraneous proteins. The purified proteins were then used to stimulate mBMDCs and the D1 cell line, representing a long-term growth factor-dependent immature myeloid DC line of splenic origin. Our results clearly demonstrate that the TgaA protein can activate in vitro dendritic cells, inducing a dose-dependent production of IL-2. A similar stimulatory effect was found with the recombinant protein CHAP, whereas LT demonstrated to activate only marginally on both these dendritic cell models, and did not show any significant ability to trigger IL-2 expression. Therefore, protein TgaA activates DCs and triggers IL-2 production through its CHAP domain. Conclusions TgaA, a protein from the human intestinal and probiotic bacterium B. bifidum MIMBb75, is a cell surface-exposed molecule that is capable to induce dendritic cell–derived IL-2 production through its CHAP domain. Thus, the CHAP domain of TgaA should be considered an active partner participating in the cross-talk mechanisms among Gram(+) pathogenic or commensal bacteria and their host.