Antibodies enantioselectivity

Figure 3.3 Antibody 14D9 catalyzes the enantioselective protonation of enol ethers.

The molecular mechanism of the enantioselective protonation reaction by antibody 14D9 was revealed by a crystal structure analysis [19[. A catalytic carboxyl group AspH 101 was found at the bottom of the catalytic pocket and found to be necessary for catalysis by mutagenesis to Asn or Ala. The mechanism or protonation involves an overall syn addition of water to the enol ether in a chiral binding pocket ensuring complete enantioselectivity (Figure 3.4). 

Like many other antibodies, the activity of antibody 14D9 is sufficient for preparative application, yet it remains modest when compared to that of enzymes. The protein is relatively difficult to produce, although a recombinant format as a fusion vdth the NusA protein was found to provide the antibody in soluble form with good activity [20]. It should be mentioned that aldolase catalytic antibodies operating by an enamine mechanism, obtained by the principle of reactive immunization mentioned above [15], represent another example of enantioselective antibodies, which have proven to be preparatively useful in organic synthesis [21]. One such aldolase antibody, antibody 38C2, is commercially available and provides a useful alternative to natural aldolases to prepare a variety of enantiomerically pure aldol products, which are otherwise difficult to prepare, allovdng applications in natural product synthesis [22]. 

An interesting case in the perspective of artificial enzymes for enantioselective synthesis is the recently described peptide dendrimer aldolases [36]. These dendrimers utilize the enamine type I aldolase mechanism, which is found in natural aldolases [37] and antibodies [21].These aldolase dendrimers, for example, L2Dl,have multiple N-terminal proline residues as found in catalytic aldolase peptides [38], and display catalytic activity in aqueous medium under conditions where the small molecule catalysts are inactive (Figure 3.8). As most enzyme models, these dendrimers remain very far from natural enzymes in terms ofboth activity and selectivity, and at present should only be considered in the perspective of fundamental studies. 

P. Dutta, C.A. Tipple, N.V. Lavrik, P.G. Datskos, H. Hofstetter, O. Hofstetter, and M.J. Sepaniak, Enantioselective sensors based on antibody-mediated nanomechanics. Anal. Chem. 75, 2342—2348 (2003). 

Fig. 9.12. High-throughput screening of enantioselective catalysts by competitive enzyme immunoassays [ 18]. The solid antibody recognizes both enantiomers, and tire hatched antibody is (S)-specific, enabling die determination of yield and ee.

Antibody Catalysis. Recent advances in biocatalysis have led to the generation of catalytic antibodies exhibiting aldolase activity by Lemer and Barbas. The antibody-catalyzed aldol addition reactions display remarkable enantioselectivity and substrate scope [18]. The requisite antibodies were produced through the process of reactive immunization wherein antibodies were raised against a [Tdiketone hapten. During the selection process, the presence of a suitably oriented lysine leads to the condensation of the -amine with the hapten. The formation of enaminone at the active site results in a molecular imprint that leads to the production of antibodies that function as aldol catalysts via a lysine-dependent class I aldolase mechanism (Eq. 8B2.12). 

Antibody-catalyzed reactions, too, can be conducted on a gram scale (Reymond, 1994) the biocatalyst was separated by dialysis in a cellulose membrane (12-14 kDa, antibody 150 kDa) and reused. The enantioselective hydrolysis of the enol ether (4) to the ketone (5) ran with an e.e. of 89-91% (Figure 18.10). 

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