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Enzyme Kinetics and Drugs as Enzyme Inhibitors
Published in Peter Grunwald, Pharmaceutical Biocatalysis, 2019
an enzyme (E)-catalyzed conversion of a substrate (S) to the product (P) via the formation of an enzyme/substrate (ES) complex (see also Schnell and Miani, 2003). Although Leonor Michaelis and Maud Leonora Menten (1913) in their famous publication assumed that the concentration of the ES-complex depends only on its equilibrium with S and E the expressions Michaelis–Menten equation and Michaelis constant have retained in the literature. The Michaelis–Menten equation can be written as
Introduction to Enzymes
Published in John C. Matthews, Fundamentals of Receptor, Enzyme, and Transport Kinetics, 2017
The equations that we will derive for the analysis of enzyme-catalyzed reactions are essentially the same equations as those we have derived for receptor-ligand interactions. In many instances they can be used in the same ways. Enzymes cannot, under most circumstances, be directly measured by their abilities to bind their substrates as we do with receptor-ligand interactions. This is because as soon as the substrate binds with the enzyme it is converted to product. It is much more straightforward to measure enzymes by following the rate of loss of substrate or the rate of formation of product. Therefore, instead of relating the amount of bound ligand to the concentration of unbound ligand, we have to relate the rate of reaction to the concentration of substrate. Only in situations where more than one substrate, or a substrate and a cofactor, are required for the reaction and one of these is omitted, can ligand binding analysis be employed to study enzyme-substrate interactions. In the next two chapters we will address the differences between the analysis of enzyme-substrate interactions and receptor-ligand interactions and derive equations that describe enzyme kinetics.
Instrumentation
Published in Clive R. Bagshaw, Biomolecular Kinetics, 2017
X-ray crystallography provides structural information at atomic resolution, although its application is usually confined to examining static structures in which protein–ligand complexes are trapped in stable states. The structures of enzyme-substrate intermediates are usually inferred from those of nonreactive substrates or transition-state analogs or obtained under conditions (pH or temperature) where catalysis is halted. Dynamic states within a crystal result can result in loss of diffraction and are not easily distinguished from static disorder. Alternatively, as discussed previously for aromatic residue in proteins, a side chain may appear static in an x-ray structure even though it is undergoing rapid flips as evidenced by NMR line shape, provided the ring spend most of its time in one plane. Crystallography is usually performed at low temperatures to minimize dynamic disorder. However, in favorable cases, the electron density recorded at ambient temperature can be interpreted in terms of two or more conformations [559,560].
Purification and characterisation of glutathione reductase from scorpionfish (scorpaena porcus) and investigation of heavy metal ions inhibition
Published in Journal of Enzyme Inhibition and Medicinal Chemistry, 2023
Metals found naturally in the environment and in water originating from natural and anthropogenic causes. Several metals and chemicals have been tested on various enzymes for their inhibitory effects19. Due to many industrial problems, heavy metal accumulation in the environment has become an important problem20. Heavy metals, which can be harmful even at low quantities, enter the body through the mouth, breathing and skin. They cannot be eliminated from the excretory tracts such as kidney, liver, intestine, lung and skin without special intervention. Consequently, almost all heavy metals accumulate in biological organisms. These metals build up in living things and cause major disorders such thyroid neurological diseases, autism and infertility. They trigger and enhance the generation of free radicals in aquatic species. Therefore, it should be kept under control in order to prevent the damage caused by free radicals resulting from heavy metals or chemicals21–25. Heavy metals and metal ions have an effect on the variables that affect enzyme-substrate and cofactor affinity. Metal ions, which induce transient depletion of GSH and inhibition of antioxidant enzymes, are known to have an effect on enzyme function26. It is also known that the glutathione reductase enzyme is highly sensitive to metal ions when the GSSG concentration is low27.
The role of kallikreins in inflammatory skin disorders and their potential as therapeutic targets
Published in Critical Reviews in Clinical Laboratory Sciences, 2021
Caitlin T. Di Paolo, Eleftherios P. Diamandis, Ioannis Prassas
Naturally derived compounds have also been identified as KLK inhibitors. Isocoumarins, which are a class of natural polyphenolic compounds, display inhibitor effects toward KLK5 and KLK7 [157]. Based on this finding, isomannide rigid scaffolds were used to devise isomannide-based peptidomimetic derivatives that improved inhibition constant (Ki) values to the submicromolar range for KLK5 [158,159]. More recently, another plant polyphenol called Brazilin was also identified with comparable Ki values to what was initially reported with the original isocoumarin compounds [160]. Fukugetin is a natural plant flavanone-flavone biflavonoid compound that also displays inhibitory activity toward the KLKs, but is unique in that it presents a mixed-type mechanism of inhibition that makes it useful for the development of an exosite-type inhibitor [161]. It is capable of binding both the free enzyme and the enzyme–substrate complex with two different affinity constants. Small natural plant-extracts called terterpenoids were also found to be selective toward only the trypsin-like KLKs [162]. 3-Acyltetramic acids are naturally occurring metabolites of terrestrial and marine organisms that exhibit a variety of biological properties such as antibacterial, antiviral, antifungal, and antihumoral activities [163]. They were found to be novel inhibitors of KLK5 and KLK7, with the nature and length of the acyl chain being crucial to the KLK5, KLK7, and trypsin inhibition activities [163].
Inhibition of catechol-O-methyltransferase by natural pentacyclic triterpenes: structure–activity relationships and kinetic mechanism
Published in Journal of Enzyme Inhibition and Medicinal Chemistry, 2021
Fang-Yuan Wang, Gui-Lin Wei, Yu-Fan Fan, Dong-Fang Zhao, Ping Wang, Li-Wei Zou, Ling Yang
The [S] and [I] represent the concentrations of substrate and inhibitor, respectively; Vmax is the maximal velocity; Km is the Michaelis constant; Ki is the inhibition constant suggesting the dissociation of the enzyme-inhibitor complex (EI); αKi is the inhibition constant when the inhibitor binds to an enzyme-substrate complex, suggesting the dissociation constant of the enzyme-substrate-inhibitor complex (ESI); the value of α equals 1, meaning the pure non-competitive inhibition; at α ≫ 1, meaning competitive inhibition; at α ≪ 1, meaning uncompetitive inhibition. The reciprocals of velocity and substrate concentrations gave the linear correlation by which Km and Vmax values can be calculated. Obtained data were presented as mean ± standard error (± SE) of three independent experiments with duplicate determinations for each assay. Statistical difference in IC50 values were determined using an unpaired two-tailed t-test, which indicated as *p < .05, **p < .01, ***p < .001.