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Synthesis of new (-)-Bestatin-based inhibitor libraries reveals a novel binding mode in the S1 pocket of the essential malaria M1 metalloaminopeptidase.
datasetposted on 2016-12-11, 22:27 authored by G Velmourougane, MB Harbut, S Dalal, Sheena McGowan, CA Oellig, James Whisstock, M Klemba, DC Greenbaum
The essential malarial PfA-M1 metalloaminopeptidase is a validated drug target that functions in the terminal stages of hemoglobin digestion. The natural product dipeptide mimetic, bestatin, is a potent inhibitor of PfA-M1 and provides an excellent scaffold for the development of novel research tools as well as more effective PfA-M1 inhibitors. Here we present a new, efficient and high yielding protocol for the synthesis of bestatin-derivatives from commercially available natural and unnatural N-Boc-D-amino acids. We developed a diverse library of bestatin derivatives with variants at the sidechain of either the Œ±-hydroxy-Œ≤-amino acid or the adjacent natural Œ±-amino acid. Surprisingly we found that large aromatic rings at the P1 position resulted in potent inhibition against PfA-M1, while small hydrophobic sidechains were favored at the P1‚Äô position. These data contrast previous studies that suggested the primary substrate specificity (S1) pocket of the PfA-M1 enzyme is unable to accommodate side-chains much larger than a P1 phenylalanine. To understand these apparently contradictory data, we determined the X-ray crystal structure of the PfA-M1 / bestatin-Tyr(OBzl) complex. The structure revealed a substantial inhibitor-induced rearrangement of the primary loop that forms the S1 pocket that permits accommodation of the bestatin-Tyr(OBzl) inhibitor. These findings are in contrast to most proteases where the S1 pocket is considered to define primary enzyme specificity through substantial rigidity. Taken together, our data provide important insights for the rational design of more potent and selective inhibitors of this enzyme, which may eventually be of therapeutic value for the treatment of malaria.