Enzyme-generated ‘ ‘reactive-intermediate

In addition to the assessment of reversible inhibition, the role played by mechanism-based inhibitors (irreversible inhibitors) provides a focus during lead development, as it can result in a more profound and prolonged effect than that suggested by the therapeutic dose or exposure. Mechanism-based inhibition (MBI) occurs as a result of the CYP generating reactive intermediates that bind to the enzyme causing irreversible loss of activity. Oxidative metabolism via that CYP is only restored upon re-synthesis of that enzyme. Three mechanisms have been reported showing how intermediate species act as mechanism-based inhibitors ... 

One remarkable feature of enzyme-catalyzed reactions is the ability to generate at enzyme sites reactive intermediates, such as carbanions, carbocations, and radicals, that normally would require strong reaction conditions to generate in chemical reactions and would be very unstable in aqueous solution. 

The acyl-enzyme can eliminate the 4-chlorine atom to generate this reactive intermediate that can then react with a nearby nucleophile such as His57 to give an alkylated acyl-enzyme derivative in which the inhibitor moiety is bound to the enzyme by two covalent bonds (Scheme 11.5). Inhibition is irreversible.59 The mechanism has been confirmed by X-ray structural analysis of protease-isocoumarin complexes. There is a cross-link between the inhibitor and the Serl95 and His57 residues of PPE.60 Human leukocyte elastase is also very efficiently inactivated.61... 

Optical fiber sensors that use enzymes can operate in the direct or indirect detection mode. In the first case, the optical properties of the reactives, intermediates or products of the biocatalyzed reaction can be monitored using the optical fibers. In the second type, an optochemical transducer generates the optical changes. 

As in the luminol case, the main role of the enhancer (EnH) seems to be related to turnover of the enzyme, generating enhancer radicals (En rad) in the process that are capable of oxidizing the acridan ester (AcH). The structure of the enhancer obviously is very important. To accelerate HRP turnover, the enhancer must on the one hand be able to rapidly react with the reactive HRP intermediates Cl and especially CII (k2 and k3 large). On the other hand, the oxidized enhancer intermediate (radical or radical cation) must be able to oxidize the acridan ester (light-generating step). This last reaction also depends on the structure of the acridan ester in a very unfavorable case, adding an enhancer for enzyme turnover could actually diminish the light production if k 4 > fct (Fig. 5), i.e., if the enhancer radical would not be able to oxidize the acridan ester. 

In general, biotransformation reactions are beneficial in that they facilitate the elimination of xenobiotics from pulmonary tissues. Sometimes, however, the enzymes convert a harmless substance into a reactive form. For example, CYP-mediated oxidation often results in the generation of more reactive intermediates. Thus, many compounds that elicit toxic injury to the lung are not intrinsically pneumotoxic but cause damage to target cells following metabolic activation. A classic example of this is the activation of benzo(a)pyrene, which is a constituent of tobacco smoke and combustion products, and is... 

This is similar to the reactive intermediate of equation 9.13 here, though, the enzyme has evolved to deal with the intermediate and is not inactivated. However, a similar but far more reactive intermediate can be generated from the reaction of the enzyme with /3-trifluoroalanine (equation 9.15), which will alkylate the enzyme and inactivate it.20... 

Biochemical and molecular toxicology consider events at the biochemical and molecular levels, including enzymes that metabolize xenobiotics, generation of reactive intermediates, interaction of xenobiotics or their metabolites with macromolecules, gene expression in metabolism and modes of action, and signaling pathways in toxic action. 

The typical one-electron redox reactions resulting from PET processes can be applied to control the generation of reactive intermediates similar to the way radical enz5unes are performing 112). To achieve an accmnulation of permanent reaction products, as is the case with most oxidoreductase enzymes and photos5mthetic systems, it is very important to provide suitable (photo)-catalytic multielectron transfer (MET) pathways. 

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