Enhancement of antigen-specific humoral immune responses and protein solubility through conjugation of bacterial flagellin, Vibrio vulnificus FlaB, to the N-terminus of porcine epidemic diarrhea virus surface protein antigen S0

Oh, Seo-ho; Kim Cho, Young-Saeng; Lee, Ho-Bin; Lee, Sang-Mok; Kim, Whee-Soo; Hong, Liang; Cho, Chong-Su; Choi, Yun-Jaie; Kang, Sang-Kee

In this study, we conjugated flagellin, Vibrio vulnificus FlaB, to the N-or C-terminus of PEDV antigen S0 and evaluated its ability to improve protein solubility and an antigen-specific humoral immune response. The ability of conjugating an immunopotentiator to an antigen to enhance the performance of the vaccine was once again confirmed. Conjugation of flagellin to S0 improved the solubility of the recombinant proteins when combined with chaperone tig. Flagellin conjugation did not improve the solubility of all other proteins; however, flagellin has been shown to be a good candidate for enhancing antigen solubility in proteins that are prone to form IBs such as S0. Conjugation of flagellin to the N-terminus of S0 enhanced the S0specific humoral immune response more than any other type of flagellin assistance. However, flagellin conjugation to the N-terminus of other antigens (e.g. HIV gp120, HPV E7) is not guaranteed to improve the antigen-specific humoral response because flagellin's preference to activate TLR5 or NLRC4/NAIP5 in its position differs depending on the conjugated antigens. Rather, we suggest that flagellin-conjugated antigens, which have an inferior ability to activate NLRC4/NAIP5, may be good candidates for inducing an increased antigenspecific humoral immune response. Furthermore, S0-induced responses were dominated by S0-specific IgG1 rather than IgG2a. Because the induction of IgG2a and IgG1 represent Th1-and Th2-type immune responses, respectively [18] . S0 seems to have an inherent ability to induce Th2-type immune response to itself (Fig. 4F) . In contrast to S0-F and F-S0, flagellin alone induced IgG2a more than IgG1, suggesting that flagellin skewed the type of immune response from Th2 to Th1. As Th2 cell-derived cytokines, such as IL-5 and IL-6, are important for the generation of IgA at mucosal site [40] , S0-F or F-S0 is expected to have an additional advantage if they are utilized as a mucosal vaccine. These findings provide crucial information for the rational design of a PED vaccine and flagellin-based immunotherapeutics.