Catalytic antibodies for complex reactions - Hapten design and the importance of screening for catalysis in the generation of catalytic antibodies for the NDA/CN reaction

Success in generating catalytic antibodies as enzyme mimics lies in the str ategic design of the transition-state analog (TSA) for the reaction of inte rest, and careful development of screening processes for the selection of a ntibodies that are catalysts. Typically, the choice of TSA structure is str aightforward, and the criterion for selection in screening is often binding of the TSA to the antibody in a microtiter-plate assay. This article empha sizes the problems of TSA design in complex reactions and the importance of selecting antibodies on the basis of catalysis as well as binding to the T SA. The target reaction is the derivatization of primary amines with naphth alene-2,3-dicarboxaldehyde (NDA) in the presence of cyanide ion. The desire d outcome is selective catalysis of formation of the fluorescent derivative in preference to nonfluorescent side-products. In the study, TSA design wa s directed toward the reaction branch leading to the fluorescent product. H ere, we describe a microtiter plate-based assay that is capable of detectin g antibodies showing catalytic activity at an early stage. Of the antibodie s selected, 36% showed no appreciable binding to any of the substrates test ed, but did show catalytic activity in derivatizing one or more of the amin o acids screened. In contrast, only two out of 77 clones that showed bindin g did not show catalysis. Thus, in this complex system, observation of bind ing is a good predictor of the presence of catalytic activity, and failure to observe binding is a poor predictor of the absence of catalytic activity .