Enzymes: Irreversible Inhibition

Andrew G McDonald, Trinity College Dublin, Dublin, Ireland
Keith F Tipton, Trinity College Dublin, Dublin, Ireland

Published online: June 2012

DOI: 10.1002/9780470015902.a0000601.pub2

Many drugs that are in clinical use work by irreversibly inhibiting specific enzymes, either in the individual being treated or in an invading organism. Enzyme inhibitors are also of value for understanding the behaviour of individual enzymes and metabolic systems. Recovery from irreversible inhibition requires the synthesis of new enzyme. Although those irreversible inhibitors that react with specific groups in the enzyme protein generally inhibit more than one enzyme, those that initially form a noncovalent complex with the enzyme, with subsequent reaction within that complex leading to the formation of a covalent bond, can show a high degree of selectivity. Kinetic studies, which are detailed in this account, can be used to distinguish the different types of inhibition and the factors that contribute to effectiveness and selectivity.

Key Concepts:

  • Irreversible inhibition cannot be reversed by the removal of the excess inhibitor from the system.
  • Recovery from reversible inhibition depends on the removal of the inhibitor from the system, whereas recovery from irreversible inhibition requires the synthesis of fresh enzyme.
  • Enzyme turnover in the tissues is a balance between the rate of its synthesis and degradation.
  • Nonspecific irreversible inhibitors can react with several groups of the same type in different proteins.
  • Specific irreversible inhibitors form an initial noncovalent complex with the enzyme before the reaction within that complex to form a covalent bond.
  • Active‐site‐directed inhibitors contain a reactive grouping attached to a substrate analogue.
  • Mechanism‐based inhibitors can be highly specific because they depend on the activity of the enzyme to form a reactive intermediate that then reacts irreversibly with the enzyme.
  • Suicide substrates behave as both substrates and irreversible enzyme inhibitors in divergent processes.
  • Dosage schedule for effective therapy with irreversible inhibitors is dependant on the half‐life of enzyme turnover.

Keywords: enzyme modification; kinetics (of inhibition); Kcat inhibitors; mechanism‐based enzyme inhibitors; suicide inhibitors; suicide substrates

Figure 1. First‐order inactivation of an enzyme. (a) Determination of the apparent first‐order rate constant (k′). (b) Dependence of k′ upon substrate concentration for a nonspecific irreversible inhibitor.
Figure 2. Irreversible inhibition of an enzyme by a nonspecific irreversible inhibitor. The rate of activity loss is given by the rate constant kA and k1′ and k′2 are the apparent rate constants for the modification of amino acid residues X and Y, which are both necessary for activity (kA=k1′+k2′). Residue Z cannot be involved since kZ′>kA, and no conclusion can be reached about residue W, which does not react.
Figure 3. Variation of the apparent first‐order rate constant for a specific irreversible inhibitor with inhibitor concentration according to eqns (17) and (19).
Figure 4. Substrate transformation in the presence of a nonspecific irreversible inhibitor (eqn (VI)). Determination of the pseudo‐first‐order rate constant (k′) and Ks according to eqn (27).
Figure 5. Substrate transformation in the presence of a specific irreversible inhibitor (eqn (VII)). Determination of the kinetic parameters according to (a) eqn (17) and (b) eqn (29).
Figure 6. Time course of substrate transformation in the presence of an irreversible inhibitor according to (a) eqn (30), also showing the loss of active enzyme, and (b) eqn (31).
Figure 7. Product formation in the presence of an irreversible inhibitor. When inhibition is complete the reaction cannot be restarted with substrate, but will restart if additional enzyme is added.
Figure 8. Kinetic analysis of reaction time courses in the presence of a specific irreversible inhibitor (eqn (VII)). (a) Variation of the apparent first‐order rate constant with substrate concentration according to eqns (28) and (29). (b) The dependence of the intercepts of the lines in (a) on the inhibitor concentration.
Figure 9. Determination of the partition ratio (r) for the inactivation of an enzyme by a suicide‐substrate, according to eqn (40).
Figure 10. Kinetic behaviour of a suicide‐substrate (eqn (IX)). (a) Analysis of the time course for enzyme inactivation analysed according to eqn (42). (b) Analysis of the time course for inhibitor disappearance analysed according to eqn (44). The meaning of the parameters M, K′ and kin is given in Table 3.
Figure 11. Recovery of enzyme activity in vivo after administration of a single dose of an irreversible inhibitor. The main curve is fitted to a simple first‐order equation. The inset shows the behaviour of the integrated form according to eqn (52).
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Enzymes: Structure and Action Mechanism
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Article Title: Enzymes: Irreversible Inhibition
 
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McDonald, Andrew G, and Tipton, Keith F(Jun 2012) Enzymes: Irreversible Inhibition. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0000601.pub2]