Specific enzyme inhibitors

Some pesticides, such as the herbicides inhibiting synthesis of amino acids in plants, are extremely selective between plants and animals and very potent. The chitin synthesis inhibitors used as insecticides are also extremely selective, because only insects and crustaceans (and fungi) make chitin. The fungicides first described are also efficient and have a high degree of selectivity, but are likely to produce effects in animals and plants because they inhibit enzymes of great importance to many types of organisms. 

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Protein engineering is now routinely used to modify protein molecules either via site-directed mutagenesis or by combinatorial methods. Factors that are Important for the stability of proteins have been studied, such as stabilization of a helices and reducing the number of conformations in the unfolded state. Combinatorial methods produce a large number of random mutants from which those with the desired properties are selected in vitro using phage display. Specific enzyme inhibitors, increased enzymatic activity and agonists of receptor molecules are examples of successful use of this method. 

A wide variety of a-tnfluoromethyl a-amino acids are readily available from the reaction of 5-fluoro-4-tnfluoromethyl-l,3 azoles with allylic alcohols [138, 139] a-Tnfluoromethyl-subsumted a-amino acids show anubactenal and antihy pertensive activity Some are highly specific enzyme inhibitors (suicide inhibitors) and may be important as bioregulators [140] Furthermore, they are interesting candidates for peptide modification... 

Nucleophilic addition reactions to A -monoprotected a-amino aldehydes 1 (Table 20) represent the beginning of the worldwide interest in peptide isosteres for the preparation of certain specific enzyme inhibitors (e.g., aspartylproteinase inhibition). Some examples of this reaction type show a relatively low diastereofacial selectivity, especially when the reactions are per-... 

Although aminoacyl-tRNA synthetases are necessary for protein synthesis in all tissues, their importance in chemical carcinogenesis is difficult to assess. Mutation induction by this pathway has been studied extensively (123), yet metabolic activation in a carcinogen-target tissue has not been demonstrated. The only exception is hepatic prolyl-tRNA synthetase activation of N-hydroxy-Trp-P-2 however, hepatic O-acetylation of this substrate also occurs to an appreciable extent (12). Further investigations involving the use of specific enzyme inhibitors would be helpful in addressing this problem. 

Designing specific enzyme inhibitors on a rational basis when one does not have a detailed three-dimensional crystal structure to which to relate is a rather sophisticated challenge. Some viable approaches to such a challenge are discussed in a review chapter by Santi and Kenyon (91) This discussion will focus on our rationale for the design of an affinity label for creatine kinase, namely N-(2,3-epoxypropyl)-N-amidinoglycine (epoxycreatine ) ... 

Two specific enzyme-inhibitor mechanisms deserve special discnssion becanse they are the basis of action for several important drags. They are the transition-state analogs and the snicide snbstrates. 

Methods. A plasma matrix-independent method has been obtained by adding specific enzyme inhibitors to all the various plasma matrices, and an insignificant matrix effect has been obtained using the following criteria ... 

The vitamin can be carried by the high-affinity receptor for low-density lipoprotein in fibroblasts [251]. It has been suggested to act in vivo as a specific enzyme inhibitor (of lipoxygenase). A vitamin-E-lipoxygenase complex in vivo could terminate the initiation of free radicals and other oxidized products. The binding of the vitamin to the enzyme is probably through the hydrophobic chain and involves one peptide [135]. 

The availability of medicinal chemistry resources can decide which approach to take in the absence of chemistry, a specific enzyme inhibitor that does not distinguish between host and parasite has little value. If chemistry is available, however, identifying a specific enzyme inhibitor can lead to the discovery of parasite-selective compounds. Random screens that search for novel structures are based on the premise that discovery of a specific inhibitor is an extremely rare event, and so the nutrient-dependent viability format is more desirable for HTS. 

You will know people (probably older men) who are on fi-blockers . These are compounds designed to block the effects of adrenaline (epinephrine) on the heart and hence to prevent heart disease. One of the best is Zeneca s tenormin. Preventing high blood pressure also prevents heart disease and certain specific enzyme inhibitors (called ACE-inhibitors ) such as Squibb s captopril work in this way. These are drugs that imitate substances naturally present in the body. 

Which of the following common drugs is not a specific enzyme inhibitor ... 

Conclusively establishing the role of potential intermediates in a biosynthetic pathway is a difficult aspect of biosynthesis. Typically, intermediates accumulate because subsequent enzymatic reactions are slow. Organisms also produce shunt metabolites that are off the main pathway and may not be further metabolized these will also accumulate. Isolation of an intermediate does not, therefore, establish intermediacy. Trapping experiments are sometimes used to overcome these problems. In the pathway A B C, where A is a known precursor of C, labeled A and non-labeled B are fed at the same time. The latter is metabolized to C and labeled B is produced from A Bis then temporarily available for isolation. An alternative approach for microbial metabolites is to mutate the organism or add specific enzyme inhibitors. This may allow intermediates to accumulate. Incorporation of a labeled, potential intermediate into a product does not prove that the intermediate lies on the main biosynthetic pathway. It may simply serve as a substrate for the enzymes involved. Only when each of the enzymes in a pathway has been isolated and characterized, and the substrate specificity determined, can the intermediates in a biosynthetic route be characterized. 

FIGURE 1.15 A synthetic Lewis blood group deteiminant/ceramide antigen (45) and the synthetic specific enzyme inhibitor immucillin H (46). 

B. Species- and Tissue-Specific Enzyme Inhibitors The Non-Classical Antifolates... 

Enzyme inhibition is one of the ways in which enzyme activity is regulated experimentally or naturally. Most therapeutic drugs function by inhibition of a specific enzyme. Inhibitor studies have contributed much of the available information about enzyme kinetics and mechanisms. In the body, some of the processes controlled by enzyme inhibition are blood coagulation (hemostasis), blood clot dissolution (fibrinolysis), complement activation, connective tissue turnover, and inflammatory reactions. 

The intact animal can be improved for experimental purposes if it is rendered abnormal in some way, by genetic malfunction, by illness, or by operation. Genetic defects, or mutations, are used widely in the study of bacterial metabolism, where they can be read ily induced, for example through irradiation by X-rays or from a radioactive source. Genetic defects frequently reveal themselves in the form of the absence of one specific enzyme, and metabolic studies with such enzymically defective preparations are of the same type as those made possible by the use of a specific enzymic inhibitor which we discussed above. Genetic defects in animals are rarer, but classic cases of the absence of specific enzymes and hence the accumulation of abnormal metabolites are provided in humans by the genetically carried diseases of phenylketonuria and alkaptonuria. In both, unusual substances are excreted in the urine, and the analysis of the reasons for their appearance has led to valuable information about the mechanism of amino acid metabolism in the body. 

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