A method for determining Control Coefficients is proposed for systems studied in vitro and applied to a model pathway. Rat liver extract, which converts glucose into glycerol 3-phosphate, was used with the addition to the incubation mixture of fructose-bisphosphate aldolase, triose-phosphate isomerase and glycerol-3-phosphate dehydrogenase as ‘auxiliary’ enzymes, which leaves all the control on the first three enzymes. The flux of the metabolic pathway was recorded by assaying NADH decay. Flux Control Coefficients (CJE) of hexokinase, glucose-6-phosphate isomerase and phosphofructokinase were calculated by titration of the system with increasing quantities of extraneous enzymes. It is shown that the summation property is fulfilled. The applicability of this procedure to study the control in any metabolic pathway is discussed. Possible relevance of the method to conditions in vivo and its limitations are considered.
Control Analysis has been carried out in the first steps of a rat liver glycolytic system. Attention has been focused on the effect of several glucose concentrations on the control, particularly regarding the role of glucokinase. From kinetic studies of the whole metabolic system we have obtained information on the flux variation under different glucose concentrations. This information together with the kinetics of glucokinase has allowed us to calculate Flux Control and Elasticity Coefficients for glucokinase and the Response Coefficient of the system with respect to glucose. The changes in of the value of Flux Control Coefficients demonstrates that in conditions of low glucose concentration, glucokinase is the main enzyme in controlling the flux through the pathway, but at high glucose concentration the control moves to phosphofructokinase. Next, we have compared our results with those obtained with the shortening and titration method, previously described (Torres, N.V., Mateo, F., Meléndez-Hevia, E. and Kacser, H., (1986) Biochem. J. 234, 169-174; Torres, N.V. and Meléndez-Hevia, E. 1991. Molec. Cell. Biochem. 101, 1-10). Furthermore, from knowledge of the enzyme kinetics of the system we have been able to build a model of the pathway that allows us computer similation of its behavior and calculation of the Flux Control Coefficient profile at different glucose concentrations. By the three methods the results correlate, supporting the use of the pathway substrate as external modulator of the metabolic system as a tool for practical application of Control Analysis.
The distribution of flux control coefficients of a glycolytic system in starved rat liver has been determined. The flux control coefficient profile in starved conditions is compared with normal fed conditions showing that in the former phosphofructokinase enhances more than 2-fold its flux control coefficient while glucokinase decreases slightly. The results also show that the starved system has a more complex structure probably because of the greater influence on the flux of the reverse substrate cycles and the synthesis and degradation fructose 2,6-bisphosphate reactions.
Control of glucose metabolism in rat liver under different glucose concentrations was studied. Flux Control Coefficients of glucokinase, glucose 6-phosphate isomerase and phosphofructokinase were determined by the 'shortening and enzyme titration' method. Results obtained show that glucose concentration in liver can play an important role in control of liver glycolysis by enhancing the Flux Control Coefficient of phosphofructokinase. Possible physiological significance of this fact is discussed.
several rat tissues Universidad de La LagunaSeparation of ATP and ADP was achieved in isotachophoresis by using malonate, inorganic phosphate (Pi), creatine phosphate and lactate as spacers, which isolate these nucleotides from other UV-absorbing products in rat tissues. By combining isotachophoresis with enzymatic end-point analysis of several rat tissues, calibration for ATP and ADP was achieved obtaining an operative calibration curve that allows the use of isotachophoresis as the only analytical technique in further assays. ATP and ADP quantities of rat skeletal and smooth muscle, heart, liver, kidney and lung were determined by isotachophoresis. Individual variation was also calculated by analyzing 30 animals in the same control conditions, muscular tissues showing the greatest values for ATP and ADP, whereas kidney and lung show the smaller values. ATP/ADP ratio is discussed as a representative parameter for describing the energy content in biological tissues, there are also experimentel reasons which imply that other product concentrations in this parameter not be included. This ATP/ADP ratio is calculated in the rat tissues studied, as well as its individual variation, obtaining the larger values for muscular tissues and the smaller for kidney. A great individual variation is found in kidney, followed by heart and skeletal muscle, and the most constant values are found in liver and smooth muscle. The results obtained in this calibration method for isotachophoresis assay suggest the use of a similar procedure in order to apply this technique for other nucleotide analysis.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.