Generation of high-affinity antibody against T cell-dependent antigen in the Ganp gene-transgenic mouse

Generation of high-affinity Ab is impaired in mice lacking germinal center-associated DNA primase (GANP) in B cells. In this study, we examined the effect of its overexpression in ganp transgenic C57BL/6 mice (Ganp ^Tg). Ganp^Tg displayed normal phenotype in B cell development, serum Ig levels, and responses against T cell-independent Ag; however, it generated the Ab with much higher affinity against nitrophenyl-chicken gammaglobulin in comparison with C57BL/6. To further examine the affinity increase, we established hybridomas producing high-affinity mAbs and compared their affinities using BIAcore. C57BL/6 generated high-affinity anti-nitrophenyl mAbs (K_D ∼ 2.50 ∼ 10^-7 M) of IgG1/λ1 and contained the V_H186.2 region with W33L mutation. Ganp^Tg generated much higher affinity (K_D > 1.57 × 10^-9 M) by usage of V_H186.2 as well as noncanonical V_H7183 regions. Ganp^Tg also generated exceptionally high-affinity anti-HIV-1 (V3 peptide) mAbs (K_D > 9.90 × 10^-11 M) with neutralizing activity. These results demonstrated that GANP is involved in V region alteration generating high-affinity Ab.