Role of Recombinant V_(H) and V_(L) Domains of Anti-Z-DNA mAb Z44 in Antigen Binding

Abstract

It has long been known that both heavy(VH) and light chain variable region(VL) of antibody are involved in antigen binding. Recently, reports for anti-DNA Abs such as Z22 and Hed-10 have revealed that VH chain is more important than VL in recoginizing antigen. However, it is not certain yet whether the VH chain alone of these specific anti-DNA antibodies has dominant role or such dominance of VH chain can be applicable to other anti-DNA Abs. To investigate this possibility in connection with the role and contribution of VL chain, experiments were performed using mAb Z44 which binds to Z-DNA but rocognizes different epitopes from Z22 which share 95% sequence homology. The recombinant proteins including Z44VH and Z44VL were produced by E.coli. After purification of the these antibodies, the affinity and specificity of recombinant Ab proteins to DNA antigens were tested. The results were as follows; Z44VH bound most strongly to Z-DNA and bound also to ssDNA, whereas Z44VL did not bind to B-DNA, ssDNA, and Z-DNA. In vitro association of Z44VH with Z44VL has increased the affinity to Z-DNA synergistically compared with Z44VVH alone. In the case of association of Z22VH with Z22VL affinity to Z-DNA has showed synergistic increment compared with Z22VH alone. However, the associations of Z44VH with Z22VL or Z22VH with Z44VL did not show any significant increments of affinity. From our findings that Z44VH alone could bind to Z-DNA or to ssDNA and that Z 44VL could not, it could be concluded that Z44VH has dominant role in recognizing the antigen. In addition, it may be suggested that Z44VL can increase the affinity of Z44VH and play some role in changing the specificity of Z44VH.