News and Publications

Designer Biocatalysts

F. Grynszpan, E. Keinan

USE OF ANTIBODIES TO DISSECT A CATALYTIC EVENT

Catalytic antibodies offer unique opportunities to examine mechanistic hypotheses and the relative importance of individual design elements in catalysis. We showed that antibodies elicited against the cyclopropenone hapten 1 catalyze ester hydrolysis with a 1000-fold enhancement in rate (Fig. 1). Because hapten 1 does not mimic the shape of the transition state yet elicits efficient catalysts, it most likely generates the necessary charge complementarity in the active site. Thus, haptenic tetrahedral geometry is a desirable feature but not a prerequisite in generating hydrolytic antibodies. This study and information on a cyclopropenone-containing protease inhibitor highlight the potential applications of the rarely used cyclopropenone functionality in the design of both haptens and enzyme inhibitors. The versatility of the antibodies to hapten 1 was indicated by their ability to catalyze the ring opening of epoxides to the corresponding chlorohydrins and the Curtius rearrangement of acyl azides.

MODELING OF PROTEINS THAT BIND RETINAL AND RETINOIC ACID

The intensive research on vitamin A and its related compounds in the past few years indicates the special importance of retinoid derivatives in a plethora of biological processes. For example, such compounds are important in transcription and in the mechanism of vision. In the latter, 11-cis-retinal covalently binds to a lysine residue within the opsin receptor. Although the protonated Schiff base of retinal with a primary amine shows a maximum at 440 nm, the corresponding absorption of the visual pigments are red-shifted approximately 150 nm.

Two lysine-containing antibodies, 38C2 and 33F12, which catalyze the aldol condensation, form a protonated Schiff base intermediate with a variety of ketones and aldehydes, imitating the behavior of the class I aldolases. We found that antibody 33F12 mimics opsin proteins in binding retinal and thereby changes its spectral properties; thus, the corresponding absorption of retinal is shifted to 510 nm. The crystal structure of the Fab fragment of 33F12 bound to retinal will provide the basis for site-directed mutagenesis to improve the rhodopsin analogy. This approach will enable us to study the isolated influence of selected parameters in vision.

ANTIBODY-CATALYZED PRODRUG ACTIVATION OF RETINOIC ACID DERIVATIVES

Retinoic acid esters that are stable in human cells were designed to be used as prodrugs. Specific esterase antibodies will be the catalysts of choice for the efficient release of retinoic acid with therapeutic purposes. Hapten 2 (Fig. 1) was designed to mimic the transition state of the selected ester hydrolysis reaction. Heterodimeric bifunctional antibodies with a subunit that specifically binds tumor cells and a second subunit that catalytically activates the prodrug compound will be used to selectively target cancer cells.

PUBLICATIONS

Grynszpan, F., Keinan, E. Use of antibodies to dissect the components of a catalytic event: The cyclopropenone hapten. Chem. Commun. 865, 1998.