Substrate utilization, competitive

Competitive Substrate Utilization. Various experiments with phenanthrene mineralization demonstrated partial inhibition with nonionic surfactants at doses less than that resulting in micellization. Such data suggest an alternative explanation for inhibition, other than surfactant effects on cell membranes and proteins. Possibly PAH-degrading microorganisms, or their competitors, utilize the surfactant as preferential substrate or carbon source. Jalvert et al. (66) made a similar conclusion about the effect of C12E4 on reductive dechlorination of hexachlorobenzene. 

Competitive inhibitors are inhibitors which have an effect on the but not on the V of an enzyme-catalysed reaction. The V is unchanged because the number of functional active sites is not altered but a greater substrate concentration is required to achieve the maximum utilization of the sites. Consequently, the for the substrate increases. Competitive inhibition may be overcome by the addition of more substrate to the enzyme reaction mixture. Competitive inhibitors often bear a structural similarity to the substrate and compete with the substrate for the active sites of the enzyme, i.e. they are isosteric. However, competitive inhibitors are not necessarily structurally analogous to the substrate, e.g. salicylate inhibition of 3-phospho-glycerate kinase, and may bind to a site distinct from the active site, e.g. L-isoleucine inhibition of threonine deaminase from Escherichia coli. The classical example of competitive inhibition is the action of malonate on succinate dehydrogenase (Figure 6.9) which advanced the elucidation of the... 

Direct and indirect competition formats, illustrated in Figure 1, are widely used for both qualitative and quantitative immunoassays. Direct competition immunoassays employ wells, tubes, beads, or membranes (supports) on to which antibodies have been coated and in which proteins such as bovine semm albumin, fish gelatin, or powdered milk have blocked nonspecific binding sites. Solutions containing analyte (test solution) and an analyte-enzyme conjugate are added, and the analyte and antibody are allowed to compete for the antibody binding sites. The system is washed, and enzyme substrates that are converted to a chromophore or fluorophore by the enzyme-tracer complex are added. Subsequent color or fluorescence development is inversely proportionate to the analyte concentration in the test solution. For this assay format, the proper orientation of the coated antibody is important, and anti-host IgG or protein A or protein G has been utilized to orient the antibody. Immunoassays developed for commercial purposes generally employ direct competition formats because of their simplicity and short assay times. The price for simplicity and short assay time is more complex development needed for a satisfactory incorporation of the label into the antibody or analyte without loss of sensitivity. 

Zinc protoporphyrin IX is a normal metabolite that is formed in trace amounts during haem biosynthesis. However, in iron deficiency or in impaired iron utilization, zinc becomes an alternative substrate for ferrochelatase and elevated levels of zinc protoporphyrin IX, which has a known low affinity for oxygen, are formed. This zinc-for-iron substitution is one of the first biochemical responses to iron depletion, and erythrocyte zinc protoporphyrin is therefore a very sensitive index of bone-marrow iron status (Labbe et ah, 1999). In addition, zinc protoporphyrin may regulate haem catabolism by acting as a competitive inhibitor of haem oxygenase, the key enzyme of the haem degradation pathway. However, it has been reported... 

Competitive inhibitors bind to specific groups in the enzyme active site to form an enzyme-inhibitor complex. The inhibitor and substrate compete for the same site, so that the substrate is prevented from binding. This is usually because the substrate and inhibitor share considerable stmctural similarity. Catalysis is diminished because a lower proportion of molecules have a bound substrate. Inhibition can be relieved by increasing the concentration of substrate. Some simple examples are shown below. Thus, sulfanilamide is an inhibitor of the enzyme that incorporates j9-aminobenzoic acid into folic acid, and has antibacterial properties by restricting folic acid biosynthesis in the bacterium (see Box 11.13). Some phenylethylamine derivatives, e.g. phenelzine, provide useful antidepressant drags by inhibiting the enzyme monoamine oxidase. The cA-isomer maleic acid is a powerful inhibitor of the enzyme that utilizes the trans-isomer fumaric acid in the Krebs cycle. 

Any compounds (other than the physiologically relevant substrate) that can serve as substrates for a particular enzyme. Alternative substrates compete with the natural substrate and with each other for access to the enzyme s active site. Thus, if one is utilizing an assay that measures the production of the true substrate, then the presence of the alternative substrate will result in competitive inhibition relative to the true substrate. 

AG 1024 has been extensively studied as an IGFR inhibitor [70] and is a substrate competitive inhibitor of this kinase [71]. AG1024 also inhibits other kinases including c-Kit [72]. Additional studies will be needed, including a direct measurement of Abl activity and possible subsequent testing against the imatinib resistant Abl point mutations, to ascertain the possible therapeutic utility of AG 1024. 

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