Pyridoxal-5 '-phosphate-dependent catalytic antibodies

Cofactors-i.e., metal ions and coenzymes-extend the catalytic scope of enzy mes and might have been among the first biological catalysts. They may be e xpected to efficiently extend the catalytic potential of antibodies. Monocl onal antibodies (MAbs) against N-alpha-phosphopyridoxyl-L-lysine were scree ned for 1) binding of 5'-phosphopyridoxyl amino acids, 2) binding of the pl anar Schiff base of pyridoxal-5'-phosphate (PLP) and amino acids, the first intermediate of all PLP-dependent reactions, and 3) catalysis of the PLP-d ependent alpha, beta-elimination reaction with beta-chloro-D/L-alanine. Ant ibody 15A9 fulfilled all criteria and was also found to catalyze the cofact or-dependent transamination reaction of hydrophobic D-amino acids and oxo a cids (k'(cat) = 0.42 min(-1) with D-alanine at 25 degrees C). No other reac tions with either D- or L-amino acids were detected. PLP markedly contribut es to catalytic efficacy-it is a 10(4) times more efficient acceptor of the amino group than pyruvate. The antibody ensures reaction specificity, ster eospecificity, and substrate specificity, and further accelerates the trans amination reaction (k'(cat(Ab))/k'(cat(PLP)) = 5 x 10(3)). The successive s creening steps simulate the selection criteria that might have been operati ve in the evolution of protein-assisted pyridoxal catalysis.