Suicide substrates, design

The efficiency of inactivation by covalent bond formation vs release of the reactive species into solution has been described by its partition ratio. The most efficient inactivators have catalytic partition ratios of 0, in which case each inhibitor molecule leads to inactivation of the enzyme. To this date, many of these inhibitors have been designed, and alternative names like suicide substrate, Trojan Horse inactivator, enzyme induced inactivator, inhibitor, and latent inactivator have been used for this class of inhibitors. A number of comprehensive reviews are available (26—32). 

The starting point for much of the work described in this article is the idea that quinone methides (QMs) are the electrophilic species that are generated from ortho-hydro-xybenzyl halides during the relatively selective modification of tryptophan residues in proteins. Therefore, a series of suicide substrates (a subtype of mechanism-based inhibitors) that produce quinone or quinonimine methides (QIMs) have been designed to inhibit enzymes. The concept of mechanism-based inhibitors was very appealing and has been widely applied. The present review will be focused on the inhibition of mammalian serine proteases and bacterial serine (3-lactamases by suicide inhibitors. These very different classes of enzymes have however an analogous step in their catalytic mechanism, the formation of an acyl-enzyme intermediate. Several studies have examined the possible use of quinone or quinonimine methides as the latent... 

More recently, Janda has described the production of a galactopyranosidase antibody in response to hapten [96]. This was designed to accommodate several features of the transition state for glycoside hydrolysis notably a flattened half-chair conformation and substantial sp2 character at the anomeric position. Some 100 clones were isolated in response to immunization with [96] and used to generate a cDNA library for display on the surface of phage (Appendix entry 7.3) (Janda et al., 1997). Rather than proceed to the normal screening for turnover, Janda then created a suicide substrate system to trap the catalytic species. 

Suicide Enzyme Inhibitors. Snicide substrates are irreversible enzyme inhibitors that bind covalently. The reactive anchoring group is catalytically activated by the enzyme itself through the enzyme-inhibitor complex. The enzyme thus produces its own inhibitor from an originally inactive compound, and is perceived to commit suicide. To design a substrate, the catalytic mechanism of the enzyme as well as the nature of the functional gronps at the enzyme active site must be known. Conversely, successful inhibition provides valuable information about the structure and mechanism of an enzyme. Componnds that form carbanions are especially usefnl in this regard. Pyridoxal phosphate-dependent enzymes form such carbanions readily becanse... 

T. M. Penning (1983). Design of suicide substrates. Trends Pharmacol. Sci. 4 212-217. 

Although detailed structural as well as mechanistic knowledge of an enzyme is desirable, it is by no means necessary in order to design a suicide substrate. This has been shown by Myers and Widlanski (1993) who have designed a simple inhibitor of human prostatic acid phosphatase (PAP), an enzyme that is believed to be involved in the regulation of androgen receptor activity in prostate cells. Since the enzyme shows a preference for hydrolysis of aryl phosphates, the 4-(fluoromethyl)-phenyl phosphate (FMPP) was prepared as a substrate that would, on hydrolysis by the... 

True enough, treatment of PAP with FMPP resulted in a time-dependent inactivation of the enzyme. Competitive inhibitors of PAP protected against inactivation. The authors suggest that FMPP represents a useful basic structure which can be incorporated into the design of more specific phosphatase inhibitors for example, the modified tyrosine 77 could be incorporated into a particular peptide to give a suicide substrate that is selective for a protein phosphatase which preferentially hydrolyses that peptide. 

The progress made at the molecular biology level in the comprehension of the mechanisms of action of /3-lactam drugs, and the mechanisms of bacterial resistance. This now allows the rational design of synthetic drugs, particularly in the field of /3-lactamase inhibitors acting as suicide substrates (see Section 2.03.12). 

Unfortunately, no useful therapeutic drug has been designed by this approach so far. Inhibiting the transaminase enzyme has no medicinal use since the enzyme is crucial to mammalian biochemistry and inhibiting it would be toxic to the host. The main use for suicide substrates has been in labelling specific enzymes. The substrates can be labelled with radioactivity and reacted with their target enzyme in order to locate the enzyme in tissue preparations. 

A group designing possible suicide substrates of AR have synthesized 17/9-hydroxy-10 -mercaptoestr-4-ene-3-one (99) and 19-mercaptoandrost-4-ene-3,17-dione (96) [212]. At variance with the 10) -amino (84) and 10)8-hydro-xy (85) estr-4-ene-3,17-dione analogues, the 17 -hydroxy-10) -mercaptoestr-... 

During the past three decades, besides the rational design of hundreds of molecules that have been synthesized and tested as suicide substrates. It also has come to light that nature Itself has known about this mechanistic mode of enzyme Inhibition and provided us with several extremely potent mechanism-based suicide Inactivators. Below are a few selected examples to demonstrate the mode of action of these Inhibitors. 

This concept of suicide substrates or enzyme inactivators has many potential implications whereby organic chemists can synthesize well-designed substrate analogues of specific enzymes. Of course, the design of new suicide enzyme inactivators involves higher and higher levels of chemical sophistication. The majority of the work done with suicide inhibitors has been on non-proteolytic enzymes, especially pyridoxal- and flavin-dependent enzymes. 

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