Promiscuity in Antibody Catalysis: Esterolytic Activity of the Decarboxylase 21D8

Abstract

Dedicated to Jack D. Dunitz on the occasion of his 80th birthdayThe high structural similarity of decarboxylase antibody 21D8 and esterase antibody 48G7 suggests that 21D8 might also possess hydrolytic activity. Kinetic investigations show that 21D8 does promote the selective hydrolysis of methyl 4-nitrophenyl carbonate and sodium 4-(acetoxy)benzenesulfonate with catalytic proficiencies (k cat /K m )/k un of ca. 10 5 m À1. The ability of 21D8 to accelerate a reaction for which it was not developed suggests that certain antibody scaffolds are intrinsically predisposed toward catalysis, a property that can be enhanced and refined during affinity maturation in response to a transition-state analog. At the same time, however, the restricted structural diversity of the immune system may ultimately limit the catalytic efficiency that can be achieved.1. Introduction. ± The immune system provides the largest combinatorial library for molecular recognition in nature. It has been estimated that the primary immunoglobulin repertoire contains ca. 10 8 different antibodies [1], and that this diversity is further expanded by several orders of magnitude through the process of somatic hypermutation [2], allowing high affinity recognition of virtually any antigen.The selectivity and diversity of the immune system have been exploited for the generation of tailored antibody receptors for many applications, including catalysis [3] [4]. The latter is achieved when a small molecule carrying chemical information about a particular reaction mechanism, such as a stable analog of a transition state, is used to induce an immune response. Catalytic antibodies, with their programmable activities, have proved to be valuable model systems for analyzing the mechanism and evolution of catalysis in proteins.In light of the many different reactions that have been catalyzed by antibodies [5], the immunoglobulin scaffold is clearly a versatile starting point for enzyme design. Nevertheless, rather than providing many distinct structural solutions to a given recognition problem, as might have been expected from the enormous sequence diversity of the primary repertoire, antibodies appear to use a relatively limited number of strategies for binding certain classes of ligand. The high degree of structural similarity in hydrolytic antibodies generated in response to aryl phosphonates and phosphonamides is a case in point [6]. Even haptens that are less obviously related can induce homologous active sites. For example, 2-(acetamido)naphthalene-1,5-disulfonate (1) and 4-nitrophenyl phosphonate 2, which both contain tetrahedral anions and hydrophobic aryl groups but are otherwise dissimilar, gave rise to antibodies that bear a striking resemblance to one another [7] (Fig. 1).

Catalytic Promiscuity in Biocatalysis: Using Old Enzymes to Form New Bonds and Follow New Pathways

“…[18] Another catalytic antibody that catalyzes decarboxylation also catalyzes ester hydrolysis. [19] In principle, changes in substrate specificity cause subtle changes in the electron distribution in the transition state and could be considered as examples of catalytic promiscuity. However, these differences are usually much smaller than the examples considered here.…”

“…Pyruvate decarboxylase, a thiamine-dependent enzyme, also catalyzes the enantioselective acyloin condensation of acetaldehyde and benzaldehyde. [19] serine hydroxymethyltransferase pyridoxal-dependent transfer of C b of serine to tetrahydropteroylglutamate threonine retroaldol reaction, decarboxylation of aminomalonate, racemization of alanine, transamination of alanine and pyruvate [26] pyruvate decarboxylase thiamine-dependent decarboxylation of pyruvate acyloin condensation of acetaldehyde and benzaldehyde [22][23][24] Angewandte Chemie U. T. Bornscheuer and R. J. Kazlauskas direct the aldimine intermediate toward a single pathway. [24] But in some cases, the aldimine can react by multiple paths.…”

Computational design of biological catalysts

“…17 Another example is the CA 21D8, which was developed to accelerate a decarboxylation reaction, and has shown hydrolytic activity. 18 These observations suggest that certain antibody scaffolds are intrinsically predisposed towards catalysis, a property that can be enhanced and refined during the affinity maturation process in response to a TSA. 18 However, as a general feature, the catalytic power of CA is never as high as the one obtained in enzymes (10 6 in CA against 10 20 in enzymes).…”

“…18 These observations suggest that certain antibody scaffolds are intrinsically predisposed towards catalysis, a property that can be enhanced and refined during the affinity maturation process in response to a TSA. 18 However, as a general feature, the catalytic power of CA is never as high as the one obtained in enzymes (10 6 in CA against 10 20 in enzymes). Their modest efficiencies appear to be a direct consequence of the simple strategy used to generate them.…”

“…It is therefore conceivable that antibody structure itself places intrinsic limitations on the kind of reactions amenable to catalysis and on attainable efficiencies. 17,18 Different strategies have been proposed to improve the catalytic efficiency of CAs. One is based on the search for a better hapten (TSA), as the TSA is never sufficiently similar to the TS.…”