Kinetic Characterization and Molecular Docking of a Novel, Potent, and Selective Slow-Binding Inhibitor of Human Cathepsin L

Penn Center for Molecular Discovery, University of Pennsylvania, 1024 Vagelos Laboratories, Philadelphia, PA 19104-6383, USA.
Molecular pharmacology (Impact Factor: 4.13). 08/2008; 74(1):34-41. DOI: 10.1124/mol.108.046219
Source: PubMed


A novel small molecule thiocarbazate (PubChem SID 26681509), a potent inhibitor of human cathepsin L (EC with an IC(50) of 56 nM, was developed after a 57,821-compound screen of the National Institutes of Health Molecular Libraries Small Molecule Repository. After a 4-h preincubation with cathepsin L, this compound became even more potent, demonstrating an IC(50) of 1.0 nM. The thiocarbazate was determined to be a slow-binding and slowly reversible competitive inhibitor. Through a transient kinetic analysis for single-step reversibility, inhibition rate constants were k(on) = 24,000 M(-1)s(-1) and k(off) = 2.2 x 10(-5) s(-1) (K(i) = 0.89 nM). Molecular docking studies were undertaken using the experimentally derived X-ray crystal structure of papain/CLIK-148 (1cvz. pdb). These studies revealed critical hydrogen bonding patterns of the thiocarbazate with key active site residues in papain. The thiocarbazate displayed 7- to 151-fold greater selectivity toward cathepsin L than papain and cathepsins B, K, V, and S with no activity against cathepsin G. The inhibitor demonstrated a lack of toxicity in human aortic endothelial cells and zebrafish. In addition, the thiocarbazate inhibited in vitro propagation of malaria parasite Plasmodium falciparum with an IC(50) of 15.4 microM and inhibited Leishmania major with an IC(50) of 12.5 microM.

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Available from: Elizabeth R Sharlow, Oct 06, 2015
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    • "diluted 100-fold into the deadenylation assay mixture [containing 7 mg poly(A)/ml] and the activity rates were monitored for 30 min further and compared to the rates of the control reaction. A rapidly reversible inhibitor should dissociate from the enzyme to restore more than approximately 90% of activity [36]. Fig. 2B summarizes the results of the preincubation/dilution assay. "
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