The long range goal of this research is to demonstrate that monoclonal antibodies (mAbs) elicited against metal complexes can be used to produce artificial metalloenzymes. The ubiquity of metalloproteins and metalloenzymes in Nature suggests that development of methods for incorporation of metal centers into antibody combining sites is a crucial step in the development of new antibody-based products. Metalloantibodies could be designed to carry out functions normally associated with metalloenzymes, and they might also be prepared to catalyze a variety of other normally metal ion dependent chemical transformations. These biotechnological products would have potential biomedical applications in pharmaceuticals, as radioisotope carriers for imaging or radiotherapy, and as scavengers for toxic substances. The focus of this proposal will be on metalloporphyrin cofactors, which will be used in """"""""cofactor/substrate analogs"""""""" to prepare catalytic metalloantibodies (metalloabzymes) that are expected to function as oxygen-atom transfer catalysts for selective oxidation of amino acids and other substances. The project will have three main phases of research concerning each of the antigens used to produce the mAbs. The first phase is the preparation of hapten-protein conjugates via synthesis of appropriately functionalized metal complexes. The second phase of work is the preparation of monoclonal antibodies and Fab antibody fragments against the haptens via hybridoma techniques or the combinatorial bacteriophage lambda vector system for expression of Fab antibody fragments in E. coli. The third phase is the characterization of the catalytic and physical properties of the metalloantibodies. The catalysis of oxidations of oligopeptides and polypeptides will be assayed for rate accelerations and selectivity for specific amino acid sequences. Physical characterization will include sequencing of the Fab fragments, molecular modeling of the hypervariable regions, spectroscopy of mAb/metal complexes (UV-Visible absorbance and circular dichroism spectrophotometry), and cyclic/differential pulse voltammetry of mAb bound metal complexes.
