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Inactivation of cysteine proteases

C P Govardhan1, R H Abeles

  • 1Vertex Pharmaceuticals, Inc., Cambridge, Massachusetts, 02254, USA.

Archives of Biochemistry and Biophysics
|June 1, 1996
PubMed
Summary
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Cysteine proteases papain and cathepsin B are inactivated by a specific Michael acceptor. The trans isomer of the inhibitor, a peptidyl-beta-chloro-alpha, beta-unsaturated ester, leads to enzyme inactivation via chloride release.

Area of Science:

  • Biochemistry
  • Enzymology
  • Medicinal Chemistry

Background:

  • Cysteine proteases are crucial enzymes involved in various biological processes.
  • Understanding their inhibition mechanisms is vital for drug development.
  • Peptidyl alpha, beta-unsaturated esters are known inhibitors of cysteine proteases.

Purpose of the Study:

  • To investigate the inactivation mechanism of cysteine proteases papain and cathepsin B by a novel Michael acceptor.
  • To elucidate the role of stereochemistry (cis vs. trans) of the inhibitor in enzyme inactivation.
  • To characterize the binding and inhibition modes of different isomers.

Main Methods:

  • Enzyme kinetics assays to determine inhibition constants and inactivation rates.
  • Stoichiometric analysis of chloride release during inactivation.

Related Experiment Videos

  • Comparison of inhibition by trans and cis isomers of the peptidyl-beta-chloro-alpha, beta-unsaturated ester.

    • Main Results:

    • The peptidyl-beta-chloro-alpha, beta-unsaturated ester effectively inactivated papain and cathepsin B.
    • Inactivation was stoichiometric with chloride release, suggesting nucleophilic attack by the active site cysteine.
    • The trans isomer caused irreversible inactivation, while the cis isomer acted as a competitive inhibitor.
    • Steric hindrance from the beta-chlorine substituent likely affects the binding of the cis isomer.

    Conclusions:

    • The study demonstrates a mechanism-based inactivation of cysteine proteases by a specific Michael acceptor.
    • The trans configuration of the inhibitor is essential for effective inactivation.
    • Stereochemistry plays a critical role in the inhibitory activity and binding mode of these compounds.