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sition state analog [70]. If applied broadly, genetic complementation could be a power-
ful tool for screening the primary immunological repertoire for more active catalysts
and for augmenting the activity of first-generation catalytic antibodies in directed evo-
lution experiments.
4.8
Future Directions
By combining programmable design with the powerful selective forces of biology, cat-
alytic antibodies merge many of the best features of synthetic and enzymatic catalysts.
Nevertheless, there is a general sense that these agents have failed to fulfill their ori-
ginal promise. While it is relatively easy to generate moderately active catalysts for
model reactions of activated substrates, production of truly efficient enzyme mimics
has proven exceptionally difficult [3]. Moreover, it has not been possible to accelerate
many reactions of interest to desirable rates. Hydrolysis of unactivated amides and
phosphate diesters are examples of such. As a consequence, visions of site-selective
proteases or restriction enzymes remain unrealized.
Reasons for these difficulties are readily enumerated [3]. Above all, the basic strategy
for producing catalytic antibodies is indirect. Tight binding to an imperfect transition
state analog, rather than catalysis, is the selection criterion that drives immunological
evolution. Even if perfect transition state analogs were available, the nanomolar affi-
nities routinely attainable during affinity maturation limit the extent to which a transi-
tion state can be stabilized relative to the ground state. Furthermore, the probability of
identifying rare but highly active clones is low because only a tiny fraction of the im-
mune response is experimentally accessible in a typical experiment.
The origins of inefficiency may also reside in the scaffold shared by all antibodies.
Immunoglobulin structural diversity appears to be far more restricted than originally
anticipated based on the vast number of germline sequences. Studies on many cat-
alytic antibodies document the frequent recurrence of basic hapten recognitionmotifs.
The commonalities observed in numerous anti-arylphosphonate antibodies [71], and
the closely related structures of the 1E9 and 39A-11 Diels-Alderases discussed above
[22], are paradigmatic. In favorable cases, such as the 1E9 antibody, the combining
pocket can be molded remarkably effectively to achieve nearly perfect shape comple-
mentarity to its ligand. In general, however, these frequently selected binding pockets
may be poorly suited to particular catalytic tasks or may represent local minima from
which it will be difficult to evolve further.
By improving transition state analogs, refining immunization and screening pro-
tocols, and developing strategies to increase the efficiency of first-generation catalytic
antibodies, it may be possible to overcome some of the hurdles in the path to enzyme-
like activities. Notably, though, the immune systemwas originally exploited as a source
of catalysts as a matter of convenience. While it is still unrivaled in biology in its ability
to fashion protein receptors to virtually any natural or synthetic molecule, new meth-
ods have since emerged for creating protein libraries based on diverse scaffolds and in
diverse formats. Phage display [72], cell surface display [73], ribosome display [74], and
