Inhibition, enzyme activity

APPLICATIONS OF GLYCAN ARRAY 15.3.1 Enzyme Activity/Inhibition Studies... 

What is fast What is slow What is relevant The relaxation time concept of Harder and Roels [148] (Fig. 28) maps typical time constants of microbial and cellular control on the level of modification of enzymes (activation, inhibition, dis/association of subunits, covalent modification or digestion) to the range of ms to s, on the level of regulation of gene expression (induction, repression or derepression of transcription) to min, on the level of population selection and evolution to days and larger units. The examples discussed below will clear up how bioengineering is facing the individual time constants. 

Not only are enzyme activities inhibited, but they can also be induced by a toxicant agent. Quantitative measures exist for a broad variety of these enzymes. Mixed-function oxidases are perhaps the best studied, with approximately 100 now identified from a variety of organisms. Activity can be measured or the synthesis of new mixed-function oxidases may be identified using antibody techniques. DNA repair enzymes can also be measured and their induction is an indication of DNA damage and associated genotoxic effects. 

Fig-1 The final NM-induced toxic effect observed in vitro is the result of multiple processes (1) interaction with proteins (formation of the protein corona, activation/inactivation of enzymes) (2) dissolution and release of toxic ions (3) production of ROS at the NMs surface (4) aggregation/agglomeration (5) diffusion and sedimentation that influence NM transport to the cell layer and the final effective concentration (6) interaction with the cell membrane and membrane receptors (activation/inhibition) (7) cell uptake (including receptor-mediated endocytosis and other uptake mechanisms) (8) interaction with intracellular enzymes (activation/inhibition) (9) production of intracellular ROS (10) activation of transcription factors and (11) binding to nucleic acids and genotoxicity, among others. Processes (1)—(5) are closely interconnected. The resulting effect observed is the result of the composite rate of all these different reactions... 

Hepatic Effects. Mild liver effects have been seen in rats and mice orally exposed to di- -butyl phthalate for acute or intermediate durations. These effects were increased liver weight, increased microsomal enzyme activity, inhibition of mitochondrial respiration, liver necrosis, and peroxisome proliferation. The lowest dose level at which effects were seen was 348 mg/kg/day. 

Figure 11.3 ROS generated by internalized NMs may induce oxidative damage to lipids, proteins, and DNA. These reactions can alter the intrinsic properties of the membrane, such as ion transport, and permeability reduce enzyme activity inhibit protein synthesis, damage DNA, and so forth, eventually resulting in cell death. Reproduced from ref. 65 with permission from John Wiley and Sons. Copyright 2013 Wiley-VCH Verlag GmbG Co. KGaA, Weinheim.

Studies in B. subtilis [151] and E. coli [148] have implicated a special role of the purine PRTases in the regulation of purine uptake and incorporation. The degree of uptake is proportional to the PRTase activity and appears to be obligatorily linked to nucleotide formation. The adenine enzyme has been found associated with membrane vesicles, and uptake by membrane preparations is completely correlated with enzyme activity. Inhibition of Ad-PRTase activity also inhibits purine adenine uptake, thus providing strong support that the observed inhibitory effects may serve to govern cellular uptake of the purines. Such a control would prevent a cell from being overloaded with excess nucleotides. 

Further degradation of the nucleosides to the purine bases by the catabolic activity of nucleoside phospho-rylase has not been detected in the assay system used. While the sum of the nucleotides and nucleo sides formed accurately reflects the enzyme activities, inhibition of the conversion of nucleotides to nucleosides by thymidine-5Hriphosphate (TTP) (5) facilitates the assay by measuring the nucleotides only. Indeed, for normal cells, the accumulation of radioactivity was found to be linear with time and protein concentration, in the nucleotide fraction only when TTP was present, while in the sum of the nucleotide and nucleoside fractions in both the presence and the absence of TTP. 

There are various modes of action of antibiotics, such as inhibition of the biosynthesis of the bacterial cell wall, action as a surface-active agent on the bacterial cell membrane, inhibition of the formation and function of bacterial enzymes by trapping metal ions, which are normally enzyme activators, inhibition of the synthesis of proteins and bacteria, action on mitochondria, and so on. 

Beyond pharmaceutical screening activity developed on aminothiazoles derivatives, some studies at the molecular level were performed. Thus 2-aminothiazole was shown to inhibit thiamine biosynthesis (941). Nrridazole (419) affects iron metabohsm (850). The dehydrase for 5-aminolevulinic acid of mouse liver is inhibited by 2-amino-4-(iS-hydroxy-ethyl)thiazole (420) (942) (Scheme 239). l-Phenyl-3-(2-thiazolyl)thiourea (421) is a dopamine fS-hydroxylase inhibitor (943). Compound 422 inhibits the enzyme activity of 3, 5 -nucleotide phosphodiesterase (944). The oxalate salt of 423, an analog of levamisole 424 (945) (Scheme 240),... 

Potassium is required for enzyme activity in a few special cases, the most widely studied example of which is the enzyme pymvate kinase. In plants it is required for protein and starch synthesis. Potassium is also involved in water and nutrient transport within and into the plant, and has a role in photosynthesis. Although sodium and potassium are similar in their inorganic chemical behavior, these ions are different in their physiological activities. In fact, their functions are often mutually antagonistic. For example, increases both the respiration rate in muscle tissue and the rate of protein synthesis, whereas inhibits both processes (42). 

If the inhibitor combines irreversibly with the enzyme—for example, by covalent attachment—the kinetic pattern seen is like that of noncompetitive inhibition, because the net effect is a loss of active enzyme. Usually, this type of inhibition can be distinguished from the noncompetitive, reversible inhibition case since the reaction of I with E (and/or ES) is not instantaneous. Instead, there is a time-dependent decrease in enzymatic activity as E + I El proceeds, and the rate of this inactivation can be followed. Also, unlike reversible inhibitions, dilution or dialysis of the enzyme inhibitor solution does not dissociate the El complex and restore enzyme activity. 

FIGURE 19.8 At high [ATP], phosphofruc-tokinase (PFK) behaves cooperatively, and the plot of enzyme activity versus [frnctose-6-phos-phate] is sigmoid. High [ATP] thus inhibits PFK, decreasing the enzyme s affinity for frnc-tose-6-phosphate. 

FIGURE 23.12 Inhibition of fructose-1,6-bisphosphatase by fructose-2,6-bisphosphate in the (a) absence and (b) presence of 25 /xM AMP. In (a) and (b), enzyme activity is plotted against substrate (fructose-1,6-bisphosphate) concentration. Concentrations of fructose-2,6-bisphosphate (in fiM) are indicated above each curve, (c) The effect of AMP (0, 10, and 25 fiM) on the inhibition of fructose-1,6-bisphosphatase by fructose-2,6-bisphos-phate. Activity was measured in the presence of 10 /xM fructose-1,6-bisphosphate. 

It is now believed that many of our useful drugs exert their beneficial action by the inhibition of enzyme activity in bacteria. Some bacteria, such as staphylococcus, require for their growth the simple organic compound poraaminobenzoic... 

Pesticide inhibition on an active enzyme has been reported, which caused enzyme activities to reduce. The collected data with and without inhibition are presented hi Table E.5.1. Determine the rate model with and without inhibitor (see Table E.5.1). Also define the type of inhibition. 

Cyclosporine A (CsA) is a water-insoluble cyclic peptide from a fungus composed of 11 amino acids. CsA binds to its cytosolic receptor cyclophilin. The CsA/cyclophilin complex reduces the activity of the protein phosphatase calcineurin. Inhibition of this enzyme activity interrupts antigen receptor-induced activation and translocation of the transcription factor NEAT to the nucleus which is essential for the induction of cytokine synthesis in T-lymphocytes. 

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