Antibodies transition-state stabilization

At the end of the review there are some examples involving catalysis by acids and bases, metal ions, micelles, amylose, catalytic antibodies, and enzymes to give the reader a feeling for how Kurz s approach may be usefully applied to other catalysts. Very few of these examples, or those involving cyclodextrins, were discussed in the original literature in the same terms. It is hoped that the present treatment will stimulate further use and exploration of the Kurz approach to analysing transition state stabilization. 

As impressive as these developments have been, chemists still have a way to go to catch up with Mother Nature . For enzymes KTS may be as low as 10 2( m, since is generally in the range 10-3 to IO m and kjku values are up to 1014 or more (Lienhard, 1973 Kraut, 1988) (see Enzymes, Section 6). Further lowering of KTS for artificial enzymes below 10 ,om will no doubt require more covalent interactions in the transition state, with better catalytic groups. Nevertheless, the transition state stabilization evident in Table 4 is comparable to that which has been achieved so far with catalytic antibodies (Section 6). 

The author has been made aware of several more references which treat transition state stabilization by metal ions in the Kurz manner (Rudakov et al., 1974 Illuminati et al., 1983 Ercolani et al., 1983 Mandolini and Masci, 1984 Galli and Mandolini, 1984) and that the Kurz approach was discussed briefly in the second edition of Hammett s famous book (Hammett, 1970). In another area, a recent special issue of Accounts of Chemical Research (vol. 26, 8, pp. 389-453 (1993)), which is devoted to Chemistry and Immunology , has several articles on catalytic antibodies. 

Clearly, the oxyanion hole is now as significant a feature of the binding site of such acyl transfer abzymes as it is already for esterases and peptidases — and not without good reason. Knossow has analysed the structures of three esterase-like catalytic antibodies, each elicited in response to the same phosphonate TSA hapten (Charbonnier et al., 1997). Catalysis for all three is accounted for by transition state stabilization and in each case there is an... 

Antibody 15C5 was able to catalyse the hydrolysis of the triester [105] with cat 2.65 x 10 3 min 1 whilst a second antibody from the same immunization programme was later found to hydrolyse the acetylcholinesterase inhibitor Paraoxon [106] with kcat = 1.95 x 10 3min-1 at 25°C (Appendix entry 6.2) (Lavey and Janda, 1996b). Antibody 3H5 showed Michaelis-Menten kinetics and was strongly inhibited by the hapten [104]. It exhibited a linear dependence of the rate of hydrolysis on hydroxide ion concentration, suggesting that 3H5 effects catalysis by transition state stabilization rather than by general acid/base catalysis. 

J.D. Stewart, S.J. Benkovic, Transition-State Stabilization as a Measure of the Efficiency of Antibody Catalysis , Nature, 375, 388 (1995)... 

A merging of chemistry and biology is essential to effectively probe the immune system for catalytic antibodies (Fig. 3). Haptens that are successful in eliciting catalytic antibodies are variations of the central theme that transition state stabilization in the antibody combining site will yield functional catalysts for a desired chemical reaction. The evolution of hapten design will be discussed further in subsequent sections. Once the hapten is selected and synthesized, it is attached to an immunogenic carrier protein, usually via an amide bond, for hyperimmunization. A preliminary screen for antibodies that bind the hapten using an enzyme-linked immunosorbent assay (ELISA) is followed by another screen for catalysis of the reaction for which the hapten... 

Perhaps the most significant contribution of the catalytic antibody field is the realization that enzyme catalysis is not simply transition state stabilization. Reactive immunization has enabled a mimic of the dynamics involved in enzyme catalysis, and an aldolase antibody that approaches enzymatic rates has been developed using this technique (59, 60). However, only a few types of reactions have been catalyzed using this hapten... 

In general, these molecules base their recognition on a specialized binding site and, similarly to enzymes, they catalyse a large variety of reactions, displaying a similar specificity, stereo specificity, kinetics and competitive inhibition, than their enzyme counterparts [453]. However, the rate of accelerations obtained by transition state stabilization still remains lower than that obtained with enzymes, i.e., 10 -10 fold [454]. In search of improved efficiency and since no stable transition state analogues can reproduce all the characteristics of the transition state analogues, new and more sophisticated alternatives have been developed to elicit catalytic antibodies [455]. These include... 

Typical antibody-catalyzed reaction rates are several hundredfold to 100,000-fold faster than the uncatalyzed reaction of the substrate. Several fundamental postulates have been proposed to explain the rate enhancements that nevertheless fall short of the enormous rate accelerations of enzymes. Is activity truly due solely to transition state stabilization by antibody-binding interactions Can additional binding interactions be built into the combining site or into the substrate molecule itself to increase the overall rate of the reaction Can new screening methods and immunological methods be developed to uncover novel catalysts with diverse activities Most important, can novel esterolytic catalysts be developed based on currently available catalytic antibody technology to efficiently hydrolyze and detoxicate cocaine ... 

J. Chen, Q. Deng, R. Wang, K. Honk, D. HUvert, Shape complementarity, binding-site dynamics, and transition state stabilization a theoretical study of Diels-Alder catalysis by antibody 1E9, Chembiochem, 2000, 1, 255-261. 

Fig. 2 (A) Antibody 43C9. elicited against a phosphonamidate hapten, catalyzes the hydrolysis of an activated amide. (B) Evidence reveals that antibody 43C9 employs a covalent catalytic mechanism. HisL91 is the nucleophile that attacks the substrate s amide carbonyl. Residue HisH35 assists in transition-state stabilization, while Trj L36 and TryH95 are probably involved in proton transfer.

Enzyme Catalyzed. The enzyme aldolases are the most important catalysts for catalyzing carbon-carbon bond formations in nature.248 A multienzyme system has also been developed for forming C-C bonds.249 Recently, an antibody was developed by Schultz and co-workers that can catalyze the retro-aldol reaction and Henry-type reactions.250 These results demonstrate that antibodies can stabilize the aldol transition state but point to the need for improved strategies for enolate formation under aqueous conditions. 

Schultz and coworkers (Jackson et a ., 1988) have generated an antibody which exhibits behaviour similar to the enzyme chorismate mutase. The enzyme catalyses the conversion of chorismate [49] to prephenate [50] as part of the shikimate pathway for the biosynthesis of aromatic amino acids in plants and micro-organisms (Haslam, 1974 Dixon and Webb, 1979). It is unusual for an enzyme in that it does not seem to employ acid-base chemistry, nucleophilic or electrophilic catalysis, metal ions, or redox chemistry. Rather, it binds the substrate and forces it into the appropriate conformation for reaction and stabilizes the transition state, without using distinct catalytic groups. 

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