Performance analysis of catalytic antibodies

In 1991, Jacobs analysed 18 examples of antibody catalysis of acyl-transfer reactions as a test of the Pauling concept, i.e. delivering catalysis by TS stabilization. The range of examples included the hydrolysis of aryl carbonates and of both aryl and alkyl esters. In some cases more than one reaction was catalysed by the same antibody, in others the same reaction was catalysed by different antibodies. 

Much earlier, Wolfenden (Westerick and Wolfenden, 1972) and Thompson (1973), established a criterion for enzyme inhibitors working as TSAs. Iliey proposed that such activity should be reflected by a linear relationship between the inhibition constant for the enzyme K and its inverse second- 

A log-log plot using K Km, /ccat and Acuncat data from the 18 separate cases of antibody catalysis exhibited a linear, albeit scattered, correlation over four orders of magnitude and with a gradient of 0.86 (Fig. 16).4 Considering the assumptions made, this value is sufficiently close to unity to suggest that the antibodies do stabilize the transition state for their respective reactions. However, even the highest A cat/A uncat value of 106 in this series (Tramontano et al., 1988) barely compares with enhancement ratios seen for weaker enzyme catalysts (Lienhard, 1973). 

4 It may also be worth mentioning here that many early estimates of Kd for the affinity of the 

A second use of this type of analysis has been presented by Stewart and Benkovic (1995). They showed that the observed rate accelerations for some 60 antibody-catalysed processes can be predicted from the ratio of equilibrium binding constants to the catalytic antibodies for the reaction substrate, Km, and for the TSA used to raise the antibody, Kt. In particular, this approach supports a rationalization of product selectivity shown by many antibody catalysts for disfavoured reactions (Section 6) and predictions of the extent of rate accelerations that may be ultimately achieved by abzymes. They also used the analysis to highlight some differences between mechanism of catalysis by enzymes and abzymes (Stewart and Benkovic, 1995). It is interesting to note that the data plotted (Fig. 17) show a high degree of scatter with a correlation coefficient for the linear fit of only 0.6 and with a slope of 0.46, very different from the theoretical slope of unity. Perhaps of greatest significance are the 

---

The catalytic performance of the MIP and appropriate controls were investigated with the hydrolysis of diphenylcarbonate. Reaction kinetics, followed by HPLC analysis of aUquots, were calculated as pseudo first-order rate constants. Rather encouragingly, the imprinted polymer showed typical Michaehs-Menten kinetics, in line with natural enzymes. The catalytic activity of the MIP (expressed as fccat/kuncat. the ratio of turnover munber of the catalysed reaction to turnover number in the absence of catalyst) was calculated to be 6900. This is markedly higher than has been reported for catalytic antibodies for carbonate hydrolysis (fccat/ uncat = 810), clearly demonstrating the great potential of MIPs for use as artificial enzymes. 

---