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New combined kinetic and thermodynamic approach to model glucose-6-phosphate dehydrogenase activity and stability

Paper ID Volume ID Publish Year Pages File Format Full-Text
18048 42710 2007 10 PDF Available
Title
New combined kinetic and thermodynamic approach to model glucose-6-phosphate dehydrogenase activity and stability
Abstract

Glucose-6-phosphate dehydrogenase (G6PD) from commercial Saccharomyces cerevisiae was concentrated by reverse micelles liquid–liquid extraction using soybean lecithin. Five successive cycles of extraction ensured a G6PD purification factor of 5.4. The kinetic and thermodynamic properties either of the concentrated fraction or the cell free extract were investigated. While the Michaelis constant for glucose-6-phosphate was shown to be almost independent of the presence of cell debris (kM = 49.3–49.4 μM), the maximum initial activity was about 16% higher in its absence, thus suggesting that impurities exerted a non-competitive-type inhibition. Moreover, the extraction seemed to slightly improve both the enzyme activity and stability. Comparison of the thermodynamic parameters of G6PD activity shows that the extraction by reverse micelles, although remarkably influenced both entropy and enthalpy contributions, had no appreciable effect on the activation free energy (ΔG* = 80.2–80.6 kJ mol−1). The enzyme was completely inactivated after 50 min at 47 °C either in the cell free extract or in the concentrated fraction. The thermodynamic parameters of G6PD thermal inactivation suggested the occurrence of two inactivation events both related to breaking of bonds responsible for the active dimer integrity: one, prevailing at low temperature, likely led to the formation of a less active dimer, while the other, prevailing at high temperature, was responsible for the formation of a totally inactive dimer or monomer.

Keywords
Glucose-6-phosphate dehydrogenase; Saccharomyces cerevisiae; Reverse micelles; Liquid–liquid extraction; Kinetic properties; Thermodynamic parametersATP, adenosine triphosphate; CF, concentrated fraction; E2A, active G6PD dimer; EI, inactive form of G6PD;
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New combined kinetic and thermodynamic approach to model glucose-6-phosphate dehydrogenase activity and stability
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 40, Issue 4, 5 March 2007, Pages 849–858
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us