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Detailed enzyme kinetics in terms of biochemical species: study of citrate synthase.

Beard DA, Vinnakota KC, Wu F - PLoS ONE (2008)

Bottom Line: The compulsory-ordered ternary catalytic mechanism for two-substrate two-product enzymes is analyzed to account for binding of inhibitors to each of the four enzyme states and to maintain the relationship between the kinetic constants and the reaction equilibrium constant.The developed quasi-steady flux expression is applied to the analysis of data from citrate synthase to determine and parameterize a kinetic scheme in terms of biochemical species, in which the effects of pH, ionic strength, and cation binding to biochemical species are explicitly accounted for in the analysis of the data.This analysis provides a mechanistic model that is consistent with the data that have been used support competing hypotheses regarding the catalytic mechanism of this enzyme.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology and Bioengineering Center and Department of Physiology, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America. dbeard@mcw.edu

ABSTRACT
The compulsory-ordered ternary catalytic mechanism for two-substrate two-product enzymes is analyzed to account for binding of inhibitors to each of the four enzyme states and to maintain the relationship between the kinetic constants and the reaction equilibrium constant. The developed quasi-steady flux expression is applied to the analysis of data from citrate synthase to determine and parameterize a kinetic scheme in terms of biochemical species, in which the effects of pH, ionic strength, and cation binding to biochemical species are explicitly accounted for in the analysis of the data. This analysis provides a mechanistic model that is consistent with the data that have been used support competing hypotheses regarding the catalytic mechanism of this enzyme.

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Related in: MedlinePlus

Fits to kinetic data from [14] on the reverse operation of kidney enzyme.Measured reverse flux as a function of concentrations of CIT and COASH was obtained from Figures 4 and 5 of [14]. Initial fluxes (µmol of COASH (or CIT) synthesized per minute per µg of enzyme) measured at the substrate concentrations indicated in the figures. All data were obtained at pH = 8.1 at 28°C. Model fits are plotted as solid lines.
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pone-0001825-g003: Fits to kinetic data from [14] on the reverse operation of kidney enzyme.Measured reverse flux as a function of concentrations of CIT and COASH was obtained from Figures 4 and 5 of [14]. Initial fluxes (µmol of COASH (or CIT) synthesized per minute per µg of enzyme) measured at the substrate concentrations indicated in the figures. All data were obtained at pH = 8.1 at 28°C. Model fits are plotted as solid lines.

Mentions: Matsuoka and Srere reported a comprehensive study on the forward and reverse kinetics of citrate synthase from rat kidney [14] that is useful in identifying the kinetic parameters for this enzyme. Data used here are plotted in Figures 2 and 3.


Detailed enzyme kinetics in terms of biochemical species: study of citrate synthase.

Beard DA, Vinnakota KC, Wu F - PLoS ONE (2008)

Fits to kinetic data from [14] on the reverse operation of kidney enzyme.Measured reverse flux as a function of concentrations of CIT and COASH was obtained from Figures 4 and 5 of [14]. Initial fluxes (µmol of COASH (or CIT) synthesized per minute per µg of enzyme) measured at the substrate concentrations indicated in the figures. All data were obtained at pH = 8.1 at 28°C. Model fits are plotted as solid lines.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2266798&req=5

pone-0001825-g003: Fits to kinetic data from [14] on the reverse operation of kidney enzyme.Measured reverse flux as a function of concentrations of CIT and COASH was obtained from Figures 4 and 5 of [14]. Initial fluxes (µmol of COASH (or CIT) synthesized per minute per µg of enzyme) measured at the substrate concentrations indicated in the figures. All data were obtained at pH = 8.1 at 28°C. Model fits are plotted as solid lines.
Mentions: Matsuoka and Srere reported a comprehensive study on the forward and reverse kinetics of citrate synthase from rat kidney [14] that is useful in identifying the kinetic parameters for this enzyme. Data used here are plotted in Figures 2 and 3.

Bottom Line: The compulsory-ordered ternary catalytic mechanism for two-substrate two-product enzymes is analyzed to account for binding of inhibitors to each of the four enzyme states and to maintain the relationship between the kinetic constants and the reaction equilibrium constant.The developed quasi-steady flux expression is applied to the analysis of data from citrate synthase to determine and parameterize a kinetic scheme in terms of biochemical species, in which the effects of pH, ionic strength, and cation binding to biochemical species are explicitly accounted for in the analysis of the data.This analysis provides a mechanistic model that is consistent with the data that have been used support competing hypotheses regarding the catalytic mechanism of this enzyme.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology and Bioengineering Center and Department of Physiology, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America. dbeard@mcw.edu

ABSTRACT
The compulsory-ordered ternary catalytic mechanism for two-substrate two-product enzymes is analyzed to account for binding of inhibitors to each of the four enzyme states and to maintain the relationship between the kinetic constants and the reaction equilibrium constant. The developed quasi-steady flux expression is applied to the analysis of data from citrate synthase to determine and parameterize a kinetic scheme in terms of biochemical species, in which the effects of pH, ionic strength, and cation binding to biochemical species are explicitly accounted for in the analysis of the data. This analysis provides a mechanistic model that is consistent with the data that have been used support competing hypotheses regarding the catalytic mechanism of this enzyme.

Show MeSH
Related in: MedlinePlus