Glucose NADPH

Fig. 15.3 Summary of strategies used to increase CoQlO production in microbes. Numbers in parentheses refer to studies listed in Tables 15.1 and 15.2. Abbreviations GLU glucose, NADPH/ NADP nicotinamide adenine dinucleotide phosphate, NADH/NAD nicotinamide adenine dinucleotide, G3P glyceraldehydes-3-phosphate, PYR pyruvate, A-CoA acetyl coenzyme A, TCA tricarboxylic acid, E4P erythrose-4-phosphate, PEP phosphoenolpyruvate, DMAPP demethylallyl diphosphate, IPP isopentenyl diphosphate, FPP farnesyl diphosphate, DPP decaprenyl diphosphate, PHBpara-hydroxybenzoate, NADH dh NADH dehydrogenase, succ. dh succinate dehydrogenase, ROS reactive oxygen species...

Two or more linked enzyme reactions can lead to a change in the concentration of NADH or NADPH that is equivalent to the concentration of the original analyte. The reference glucose measurement using hexokinase [9001-51-8] and glucose-6-phosphate dehydrogenase [9001-40-5] is an example ... 

Utilization of Glucose-6-P Depends on the Cell s Need for ATP, NADPH, and Ribose-5-P... 

Figure 7-10. Coupled enzyme assay for hexokinase activity. The production of glucose 6-phosphate by hexokinase is coupled to the oxidation of this product by glucose-6-phosphate dehydrogenase in the presence of added enzyme and NADP". When an excess of glucose-6-phosphate dehydrogenase is present, the rate of formation of NADPH, which can be measured at 340 nm, is governed by the rate of formation of glucose 6-phosphate by hexokinase.

The pentose phosphate pathway is an alternative route for the metabolism of glucose. It does not generate ATP but has two major functions (1) The formation of NADPH for synthesis of fatty acids and steroids and (2) the synthesis of ribose for nucleotide and nucleic acid formation. Glucose, fructose, and galactose are the main hexoses absorbed from the gastrointestinal tract, derived principally from dietary starch, sucrose, and lactose, respectively. Fructose and galactose are converted to glucose, mainly in the liver. 

Genetic deficiency of glucose-6-phosphate dehydrogenase, with consequent impairment of the generation of NADPH, is common in populations of Mediterranean and Afro-Caribbean origin. The defect is manifested as red cell hemolysis (hemolytic anemia) when susceptible individuals are subjected to oxidants, such as the an-timalarial primaquine, aspirin, or sulfonamides or when... 

The pentose phosphate pathway, present in the cytosol, can account for the complete oxidation of glucose, producing NADPH and COj but not ATP. 

The pentose phosphate pathway is operative in the RBC (it metabolizes about 5-10% of the total flux of glucose) and produces NADPH hemolytic anemia due to a deficiency of the activity of glucose-6-phosphate dehydrogenase is common. 

Another common procedure which is used for glucose assay is the hexokinase procedure in which the glucose is phosphory-lated by means of ATP and then dehydrogenated with glucose-6-phosphate dehydrogenase measuring the abosrption of NADPH... 

Based on the difference in the rate of NADPH production with both glucose-6-phosphate and 6-phosphogluconolactone as substrates and the rate of NADPH production with only 6-phos-phogluconolactone as its substrate (107). 

An excess of PGD is added to the reaction mixture containing glucose 6-phosphate and NADP to assure that 2 moles of NADPH are produced per mole of glucose-6-phosphate oxidized (109). This method has been improved by Nicholson et. al (110) to make... 

Human CYPs are multicomponent enzyme systems, requiring at a minimum the CYP enzyme component and a reductase component to be functional. The reductase requires a reduced nicotinamide cofactor, typically NADPH, and this cofactor must be regenerated to provide a steady supply of reducing equivalents for the reductase. Regeneration is accomplished with a separate substrate and enzyme. Glucose-6-phosphate and glucose-6-phosphate dehydrogenase have been widely used for this purpose. The overall complexity of the reaction mixtures and their cost have been barriers to the widespread use of recombinant human CYPs for metabolite synthesis in the past. 

The importance of having adequate supplies of NADPH for the regeneration of these various enzymes cannot be over emphasized. In normal situations this cofactor can be adequately provided by the reductive pentose phosphate pathway. Monitoring the activity of the pentose phosphate pathway has been proposed as a unique way to study the metabolic response to oxidative stress, since the glutathione peroxidase activity is coupled via glutathione reductase to the enzyme glucose-6-phosphate dehydrogenase (Ben Yoseph et ah, 1994). 

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