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Closer examination of the active site shows that the residues involved in catalysis (most notably the
residues analogous to Asp43, and Tyr48, and Lys77 in ALR2) are structurally conserved among each of
these proteins (Figure 8), suggesting that the mechanism is also conserved throughout the family. Many
of the residues found in the binding site (defined as those making contact with the zopolrestat in the
ternary complex) are also largely conserved with the exception of a number of residues in the carboxy
terminus of the protein. This is indeed where the most structure variation appears to be concentrated
among the proteins. These residues compose a loop that is the same loop that shifts upon binding of
zopolrestat in ALR2. Inhibitors with improved specificity will very likely take advantage of the subtle
structural differences that are introduced by the variation in sequences in this area.
VI. Future Design of Aldose Reductase Inhibitors
The availability of structural data for ALR2 in its holoenzyme and different ternary forms is likely to
lead to improvements in the affinity of future generations of inhibitors. As the architecture and plasticity
of the binding site are better understood, increasingly potent inhibitors may be designed to occupy it.
Although these structures provide a positive target for drug design, there are a number of negative
targets. Increased in vivo potency is likely to be derived from the specific inhibition of ALR2 that would
entail the avoidance of other members of the aldo-keto reductase family. Determination of the structures
of other members of the family may increase the specificity of compounds by providing structures of
targets to avoid. While the incorporation of the negative targets in the drug-design process relies on the
determination of other structures and is likely to be complicated, conventional computational techniques
may be applied to the problem of the positive target. Two such methods that may hold promise are
docking [43] and computational thermodynamic perturbation [44].
A. Inhibitor Docking to the Enzyme
Our initial efforts to exploit the ALR2 holoenzyme structure for drug design utilized the program DOCK
[43]. This program is capable of finding depressions on the surface of the enzyme that could serve as
binding sites for substrates or inhibitors. Once the correct area is defined, the program rotates structures
of candidate compounds within this space and scores each compound based upon its steric
complementarity with the binding site. However, the program does not include the potential polar
interactions between the inhibitor and protein when scoring. The search was further constrained by the
inability to include conformational variations both in the test compounds as well as the protein, due to
computational limitations.
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