Strong and specific peroxidase activity with an antibody L chain - Porphyrin Fe(III) complex

We attempt to generate monoclonal antibodies as host proteins for the functional molecular porphyrin, and thus engineer a new type of catalytic antibody. TCPP (meso-tetrakis (4-carboxyphenyl) porphyin) was chemically synthesized and Balb/c mice were immunized using TCPP as a hapten. Two hybridoma cells (03-1, 13-1), which produce monoclonal antibody against TCPP, were obtained. One of the monoclonal antibodies, Mab 03-1, exhibited enhanced peroxidase activity when TBP Fe(III) was incorporated. Genes for both H and L chains of monoclonal antibodies were cloned, sequenced and overexpressed using E. coli as a host, ELISA and fluorescence quenching method show that the antibody L chains from both Mab 03-1 and Mab 13-1 have specific strong interaction with TCPP. Furthermore, a TCPP Fe(III) complex with the L chain from Mab 13-1 exhibits much higher peroxidase activity than TCPP Fe(III) alone. The enzyme activity was detectable with pyrogallol and ABTS(2,2-azinobis 3-ethylbenzthiazolin-6-sulfonic acid) but not with catechol. This result provides a new concept of catalytic antibody using monoclonal antibody or antibody fragment as a host carrier protein for a functional molecule.