[Show abstract][Hide abstract] ABSTRACT: Screening of inhibitory Ab1 antibodies is a critical step for producing catalytic antibodies in the anti-idiotypic approach.
However, the incompatible surface of the active site of the enzyme and the antigen-binding site of heterotetrameric conventional
antibodies become the limiting step. Because camelid-derived nanobodies possess the potential to preferentially bind to the
active site of enzymes due to their small size and long CDR3, we have developed a novel approach to produce antibodies with
alliinase activities by exploiting the molecular mimicry of camel nanobodies. By screening the camelid-derived variable region
of the heavy chain cDNA phage display library with alliinase, we obtained an inhibitory nanobody VHHA4 that recognizes the
active site. Further screening with VHHA4 from the same variable domain of the heavy chain of a heavy-chain antibody library
led to a higher incidence of anti-idiotypic Ab2 abzymes with alliinase activities. One of the abzymes, VHHC10, showed the
highest activity that can be inhibited by Ab1 VHHA4 and alliinase competitive inhibitor penicillamine and significantly suppressed
the B16 tumor cell growth in the presence of alliin in vitro. The results highlight the feasibility of producing abzymes via anti-idiotypic nanobody approach.