Biomedical Engineering Reference
In-Depth Information
PfA-M17) in a similar fashion, but PfA-M17 also contributes a second cation
that may account for the increased potency of the inhibitors against this
enzyme.
7.3.1.1 Bestatin
Bestatin, the Phe-Leu dipeptide analog, is an inhibitor of multiple clans of
MAPs, including the M1 and M17 families. Bestatin is a potent inhibitor of
PfMAP activity in malaria cell extracts and blocks the growth of Pf parasites in
vitro.
31
Bestatin inhibits the enzymatic activity of recombinant PfA-M1 and
PfA-M17 at the same concentration.
30,31
Bestatin resembles a Phe-Leu dipep-
tide substrate; however, the first residue contains an a-hydroxy b-amino acid
that, along with the neighboring carbonyl, co-ordinates the catalytic zinc atom
resulting in a competitive active-site-directed inhibitor. The interactions of the
enzymes with bestatin are noncovalent and reversible, and bestatin is thought
to act as a slow, tight-binding inhibitor, involving the initial formation of a low-
anity complex followed by a slow conformational change leading to a high-
anity complex.
46
Thus, bestatin represents an ideal scaffold for inhibitor
development due to its family specificity, potency in the low-nanomolar to
micromolar range, synthetic tractability, and potential for expansion through
variation of the amino acid side chains in its core structure.
In both bestatin-bound enzyme structures (3KR4.pdb, 3EBH.pdb), the
bestatin 'scaffold' of a hydroxyl group (O2), central nitrogen (N2), and car-
bonyl group (O
3
) is responsible for mediating the interactions with the active-
site metals.
30,42
Chelation of these metals results in the inhibitory effect on the
enzyme. In PfA-M17, bestatin forms hydrogen bonds with 12 residues in the
active site (predominantly with metal coordinating residues), four more than
PfA-M1, again a result of the second metal ion. In the PfA-M17 enzymes, the
bestatin compound buries a larger surface area than in PfA-M1, and the P1 Phe
side chain is well packed into a narrow hydrophobic pocket forming six van der
Waals interactions. Interestingly, PfA-M1 forms a similar number of packing
interactions, but the S1 pocket of PfA-M1 was significantly larger and provides
a more flexible geometry with residues noted to move upon inhibitor binding.
The P1
0
Leu moiety for in both enzyme structures had few interactions.
7.3.1.2 Phosphinate Dipeptide Analogs
Skinner-Adams et al.
46
demonstrated that synthetic phosphinate dipeptide
analogs that inhibit metallo-aminopeptidases prevented the growth of wild-
type and the chloroquine-resistant parasites in culture. Two compounds,
hPheP[CH
2
]Phe (Compound 4, Co4) and hPheP[Ch
2
]Tyr (Compound 5), had
inhibitory constants that were superior to bestatin. The phosphinate
hPheP[CH
2
]Gly had little activity against the recombinant PfA-M1 and PfA-
M17, and did not exhibit anti-malaria activity in vitro, thus demonstrating the
importance of the hydrophobic P1
0
group. In the ultimate test of an inhibitor,
