Estimation of the overall kinetic parameters of enzyme inactivation using an isoconversional method

Abstract

Please cite this article as: D. Oancea, Alexandrina Stuparu, Madalina Nita, Mihaela Puiu, Adina Raducan, Estimation of the overall kinetic parameters of enzyme inactivation using an isoconversional method, Biophysical Chemistry (2008Chemistry ( ), doi: 10.1016Chemistry ( /j.bpc.2008 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT ABSTRACTAn isoconversional method is proposed in order to calculate the kinetic parameters of enzyme inactivation. The method provides an efficient and low-cost procedure to describe both operational and thermal inactivation. Unlike the ordinary kinetic assays performed at constant enzyme concentration and at various substrate concentrations, the isoconversional method requires several extended kinetic curves for constant initial substrate concentration and different enzyme concentrations. The procedure was tested and validated using simulated data obtained for several kinetic models frequently discussed in the literature. After the validation, the isoconversional method was used for the investigation of the thermoinactivation of urease during urea hydrolysis in self buffered medium and the operational inactivation (destructive oxidation by excess peroxide) of catalase at high concentration of hydrogen peroxide. The results showed that the isoconversional method gives good results of global inactivation constant for both simple and more complex models.

Isoconversional Kinetics of Polymers: The Decade Past

“…Oancea at al . have developed a new application of an isoconversional method for determining kinetic parameters of the thermal and operational inactivation of enzymes.…”

Assessing operational inactivation of horseradish peroxidase during Brilliant Blue G decolorization

“…This method was previously used to detect the suicide inactivation of catalase at high substrate concentration, thermo-inactivation of catalase and urease. 25 For HRP, the reported inactivation rate constants was around 10 -3 s -1 for guaiacol and capsaicin 26 and 10 -2 s -1 for 2-aminophenol. 27 In the case of BB oxidation, the isoconversional times were calculated for two settled values of BB concentration, corresponding to an initial part of reaction when 15% of BB is decomposed and at moderate substrate conversion, when 50% of BB is decomposed.…”

“…24 The examination of extended progress curves (concentration of either substrate or product vs. time) performed at settled initial concentration of substrates and different initial enzyme concentration, allows the estimation of the overall inactivation constant using an isoconversional method. 25 The method involves the determination of the time values (t [ ] ( )…”

Thermal Analysis

“…The activation energies of gel melting have been positive and decrease throughout the process, whereas the numerical values of the activation energy have been found to depend systematically on the conditions of preparation such as the time and/or the temperature of gelation and the concentration of the gelatin solution . An isoconversional method has been applied by Oancea et al to evaluate the kinetics of enzyme inactivation. It has been concluded that the method yields good estimates for the global inactivation constant.…”