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protein kinase core is essential. Yet to predict the region of the structure of the inhibitor that will form
these strong Watson-Crick hydrogen bonds remains difficult.
Page 225
The use of the 6-amino group and N1 nitrogen of the purine base to model the pharmacophore, as seen
with staurosporine, is not possible in two other adenine-type inhibitors. Both isopentenyl adenine, a
nonspecific inhibitor of protein kinases, and olomoucin, a more specific inhibitor of Ser/Thr protein
kinases, are modified only at the 6-amino group position. Thus, bidentate hydrogen-bond formation as
seen in the ATP purine base is not possible. Furthermore, there are inhibitors that do not contain the
chemical structure of adenine, for example des-chloro-flavopyridol, a potent inhibitor of cdc-2 cell cycle
kinase.
In crystallographic analysis of the binding of three inhibitors, olomoucin (OLO), isopentenyl adenine
(ISO) and des-chloro-flavopyridol (DFP) to inactive CDK-2 cell cycle protein kinase, Kim and coworkers
[26] have provided additional insight into the binding of adenine-and nonadenine-based
inhibitors. Inhibitors with purine rings (OLO and ISO) bind in relatively the same area of the binding
cleft as the adenine ring of ATP. Relative orientation of each purine ring with respect to the protein is
different for all three ligands. This is most likely due to the fact that the 6-amino group of adenine in
ATP is replaced by an isopentenylamino group in ISO and by the bulky benzylamino group in OLO. In
the case of the third inhibitor, which is not an adenine derivative, the benzopyran ring occupies
approximately the same region as the purine ring of ATP. The two ring systems overlap in the same
plane but benzopyran is rotated about 60 degrees relative to the adenine of ATP. In this orientation, two
strong bidendate hydrogen bond are formed with the oxygens in the 4th and 5th positions of the
inhibitor. Furthermore, these bonds are the same ones formed by the 6-amino group and N1 nitrogen of
the adenine ring.
Crystallographic analysis has shown that both the substrate and ATP-binding clefts are structurally
conserved yet differ in the surface charges between individual protein kinases. The structural template of
the protein kinase family as discovered in the structure solution of cAPK predicts these differences.
Template modeling provides a rational basis for the design of specific inhibitors for protein kinases
based on the ATP binding site [25]. This is the first significant step in the design of specific inhibitors
targeted at the ATP site.
Recent work has now shown the conformational diversity of inhibitors binding in the interdomain ATPbinding
cleft [26]. Although the residues of the protein kinase catalytic core that form the bidentate
donor-acceptor bond with inhibitors are identical throughout different structures, the residues of the
inhibitors vary greatly. All inhibitors use this common bidentate bond yet the specificity lies in several
other bonds formed between the inhibitor and specific regions of the individual protein kinases.
Furthermore, it is difficult to model
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