Quiescent affinity labels

However, the conversion of omeprazole to the active sulphenamide does not result in formation of a reversible enzyme inhibitor, but rather results in in situ formation of a powerful affinity label. Hence we can consider omeprazole to be a unique example of quiescent affinity labeling in which selectivity results from the unique environment of the target enzyme. 

Rhinoviri are the causal agents of common colds in humans. Viral replication and maturation is dependent on proteolytic processing of a viral polyprotein by a cysteine protease known as 3C protease. The active-site cysteine in 3C protease 

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Figure 8.2 Mechanisms of irreversible enzyme inactivation. (A) Nonspecific affinity labeling, (B) quiescent affinity labeling, and (C) mechanism-based inactivation.

More recently attempts to generate highly selective quiescent affinity labels have been made for a number of protease and kinase targets. As examples, inhibitors of the Rhinovirus 3C protease (Mathews et al 1999) and of the epidermal growth factor receptors (Boschelli, 2002), both incorporating Michael acceptors to covalently inactivate cysteine residues in their target enzymes (Lowry and Richardson, 1981 Figure 8.6), have entered human clinical trials for the treatment of rhinovirus infection and cancer, respectively. 

Table 8.1 Some examples of quiescent affinity labels of clinical interest...

The value of kohs for this type of mechanism is a saturable function of [/], as was the case for quiescent affinity labels (vide supra). For this mechanism, kima (as defined above) is a complex mixture of rate constants ... 

Hence, as we saw with quiescent affinity labels, we must treat KY as a kinetic constant, not an equilibrium constant. Only in the situation that both k3 and k4 are very large (i.e., rapid equilibrium) and k5 is rate-limiting, can we equate A) with A). If, for example, k7 is even partially rate-limiting, A) > K hence the two constants have different meanings. 

Despite the mechanistic differences in the definitions of kimcl and A) between quiescent affinity labels and mechanism-based inactivators, the dependence of kohs on [/] is the same for both mechanism. Hence we cannot determine whether or not a compound is acting as a mechanism-based inhibitor, based merely on this two-step kinetic behavior. However, there is a set of distinguishing features of mechanism-based inactivation that are experimentally testable. Compounds that display all of these features can be safely defined as mechanism-based inactivators. 

Aspirin, acetylsalicylic acid Cyclooxygenase-2 "Quiescent" affinity label... 

Suicide substrates and quiescent affinity labels, unlike the other types of inhibitors discussed in this chapter, form covalent bonds with active site nucleophiles and thereby irreversibly inactivate their target enzymes. A suicide substrate,191 also described by Silverman in a comprehensive review1101 as a mechanism-based inactivator, is a molecule that resembles its target enzyme s true substrate but contains a latent (relatively unreactive) electrophile. When the target enzyme attempts to turn over the... 

One approach that avoids the release of potent electrophiles into the general cellular milieu is use of the quiescent affinity label.1131 These evolved from the classic (non-quiescent ) affinity labels such as L-l-chloro-3-tosylamido-4-phenyl-2-butanone (Fig. 7a)... 

Figure 7. Two examples of irreversible inactivators that are not suicide substrates a) TPCK, a classic" affinity label of the serine protease chymotrypsin, b) ZFK-CH2-mesitoate, a quiescent" affinity label of the cysteine protease cathepsin B, and c) the kinetic scheme for both forms of affinity label-inactivation.

A. Krantz, Peptidyl (Acyfoxy)methanes as Quiescent Affinity Labels for Cysteine Proteases , Methods Enzymol. 1994,244, 656-671... 

CONTENTS Introduction to the Series An Editor s Foreword, Albert Padwa. Preface, Bruce E. Maryanoff and Cynthia A. Maryanoff. Computer Assisted Molecular Design Related to the Protein Kinase C Receptor, Paul A. Wenderand Cynthia M. Cribbs. Chemistry and Biology of the Immunosuppressant (-)-FK-506, Ichiro Shinkai and Nolan H. Sigal. The Development of Ketorolac Impact on Pyrrole Chemistry and on Pain Therapy, Joseph M. Muchowski. Application of Silicon Chemistry in the Corticosteroid Field, Douglas A. Livingston. Hu-perzine A-A Possible Lead Structure in the Treatment of Alzheimers Disease, Alan P. Kozikowski, X.C, Tang and Israel Hanin. Mechanism-Based-Dual-Action Cephalosporins, Harry A. Albrecht and James G. Christenson. Some Thoughts on Enzyme Inhibitors and the Quiescent Affinity Label Concept, Mien Krantz Index. 

The best definition of the third type of irreversible enzyme inhibitor, mechanism-based inactivators, is provided by Dr Richard Silverman, a leading authority on the subject. A mechanism-based inactivator (sometimes, much to my dismay, called a suicide substrate), he writes, is an unreactive compound that has a structural similarity to a substrate or product for an enzyme. Once at the active site of the enzyme, it is converted into a species that generally forms a covalent bond to the enzyme, producing inactivation. Although both quiescent affinity labels and mechanism-based inactivators can be mistaken for... 

This concern isn t entirely theoretical penicillins, quiescent affinity label as mentioned above, cause allergies (mostly skin rashes and hives) in about 2% of the patients who take them, and even potentially fatal anaphylaxis in a few, and toxicity is believed to proceed through this sort of mechanism. With the current atmosphere about drug safety, it s no wonder that many companies will only consider working on irreversible inhibitors where the benefits clearly outweigh the risks as in the case of cancer, in the context of tool compounds, or perhaps not at all. Had penicillin been discovered today quite a few companies might choose not to develop it. 

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