Chemistry Reference
In-Depth Information
10
3
- and 10
5
-fold over background. Although their activity is typically many orders of
magnitude below that of comparable enzymes, they generally exhibit programmable
and enzyme-like regio- and stereoselectivity.
4.3
Evolution of Binding Affinity and Catalytic Efficiency
Acyl transfer has been the most thoroughly investigated process with respect to anti-
body catalysis. Attention paid to such systems reflects the broad utility of hydrolytic
enzymes, on the one hand, and the availability of excellent transition state analogs, on
the other.
Phosphonates and phosphonamidates, which mimic the tetrahedral geometry and
anionic character of the transition state for hydrolysis, have proven especially reliable
as haptens. Antibodies generated against such compounds readily promote the clea-
vage of esters and, in a few cases, amides. High levels of stereospecificity are attainable
even at chiral centers remote from the reaction site [11].
Antibody 48G7, elicited with p-nitrophenyl phosphonate
1
, is a representative ester-
ase [12]. It accelerates the hydrolysis of activated ester
2a
and carbonate
2b
by factors of
>
10
4
(Scheme 4.1). Detailed study of its structure and immunological origins has pro-
vided valuable insight into the mechanism and evolution of catalysis.
As programmed by the phosphonate hapten, 48G7 is a relatively simple catalyst that
facilitates direct hydroxide attack on the scissile carbonyl of its ester substrate. The
antibody-hapten complex shows
1
bound in an extended conformation with the
aryl leaving group buried deeply in a hydrophobic cleft [13]. The negatively charged
phosphonate sits near the entrance of the pocket, where it forms hydrogen bonds with
the side chains of Tyr
H33
, His
H35
, Arg
L96
, and the backbone amide of Tyr
H96
. This col-
lection of polar residues is reminiscent of the oxyanion hole in serine proteases, and
presumably stabilizes the oxyanionic transition states of the hydrolysis reaction
through analogous hydrogen-bonding and ionic interactions.
Scheme 4.1 Hydrolysis of activated aryl esters 2a and carbonates 2b
proceeds via an anionic, tetrahedral intermediate (in square brackets).
Hydrolytic antibody 48G7 was elicited with an aryl phosphonate deri-
vative 1 that mimics this high energy species and its flanking transition
states.
