Analysis of progress curves for enzyme-catalyzed reactions: application to unstable enzymes, coupled reactions and transient-state kinetics

Duggleby R.G. (1994) Analysis of progress curves for enzyme-catalyzed reactions: application to unstable enzymes, coupled reactions and transient-state kinetics. Biochimica et Biophysica Acta (BBA)/Protein Structure and Molecular, 1205 2: 268-274. doi:10.1016/0167-4838(94)90244-5


Author Duggleby R.G.
Title Analysis of progress curves for enzyme-catalyzed reactions: application to unstable enzymes, coupled reactions and transient-state kinetics
Journal name Biochimica et Biophysica Acta (BBA)/Protein Structure and Molecular
ISSN 0167-4838
Publication date 1994-04-13
Sub-type Article (original research)
DOI 10.1016/0167-4838(94)90244-5
Volume 1205
Issue 2
Start page 268
End page 274
Total pages 7
Subject 1303 Specialist Studies in Education
1304 Biophysics
1312 Molecular Biology
1315 Structural Biology
Abstract There are several advantages to the use of progress curves to analyze the kinetic properties of enzymes but most studies still rely on rate measurements. One of the reasons for this may be that progress curve analysis relies on the enzyme and the reactants being completely stable under assay conditions. Here a method is described that relaxes this requirement and allows progress curve analysis to be applied to unstable enzymes. The procedure is based on a combination of numerical integration and non-linear regression to fit rate equations to the progress curve data. The analysis is verified using simulated data and illustrated by application to the reaction catalyzed by alkaline phosphatase, measured in the presence of 10 mM EGTA where it has a half-life of 3 1 2 min. The method may also be applied to other experimental systems where the development over time reveals important properties but where an analytical solution of the underlying model is not known. This extension is illustrated by two systems: the coupled reactions catalyzed by pyruvate kinase and lactate dehydrogenase under conditions where both enzymes have similar activity; and the transient-state kinetics of the reaction catalyzed by glutamate dehydrogenase.
Keyword Coupled reaction
Enzyme
Enzyme-catalyzed reaction
Progress curve
Transient-state kinetics
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status Unknown

Document type: Journal Article
Sub-type: Article (original research)
Collection: Scopus Import
 
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