Inactivation rate constant

Dekker et al. [170] studied the extraction process of a-amylase in a TOMAC/isooctane reverse micellar system in terms of the distribution coefficients, mass transfer coefficient, inactivation rate constants, phase ratio, and residence time during the forward and backward extractions. They derived different equations for the concentration of active enzyme in all phases as a function of time. It was also shown that the inactivation took place predominantly in the first aqueous phase due to complex formation between enzyme and surfactant. In order to minimize the extent of enzyme inactivation, the steady state enzyme concentration should be kept as low as possible in the first aqueous phase. This can be achieved by a high mass transfer rate and a high distribution coefficient of the enzyme between reverse micellar and aqueous phases. The effect of mass transfer coefficient during forward extraction on the recovery of a-amylase was simulated for two values of the distribution coefficient. These model predictions were verified experimentally by changing the distribution coefficient (by adding... 

Dekker et al. [170] have also shown that the steady state experimental data of the extraction and the observed dynamic behavior of the extraction are in good agreement with the model predictions. This model offers the opportunity to predict the effect of changes, both in the process conditions (effect of residence time and mass transfer coefficient) and in the composition of the aqueous and reverse micellar phase (effect of inactivation rate constant and distribution coefficient) on the extraction efficiency. A shorter residence time in the extractors, in combination with an increase in mass transfer rate, will give improvement in the yield of active enzyme in the second aqueous phase and will further reduce the surfactant loss. They have suggested that the use of centrifugal separators or extractors might be valuable in this respect. 

Disraption of side chain interactions results in denaturation of the protein, and the rate of inactivation follows first order kinetics in the simplest cases. The plot of the logarithm of the remaining activity (In a ) versus time gives a straight line, the slope of which is the negative value of the inactivation rate constant. 

Alternatively, if the individual data have not been normalized for the spontaneous loss of CYP activity, such a correction can be applied at this step. In this approach, the control activity in the above equation is always the zero-minute control for the solvent, rather than the solvent control at each time point. The apparent inactivation rate constant for the vehicle control, Obs[i]=0 is then accounted for in the nonlinear regression according to the following equation (124) ... 

The experiment is repeated at several different temperatures to generate a wide range of inactivation rate constants. 

Irreversible inactivation of L. mingrelica luciferase at 37°C. Thermal inactivation of the WT luciferase and its mutants was investigated at 37°C at various enzyme concentrations (10 6-10"8 mol/L). Two-exponential time-curves of thermoinactivation corresponding to the fast and the slow phases of inactivation were observed. The fast hi) and slow (k2) inactivation rate constants of the luciferase were shown to be dependent on the enzyme concentration. This phenomenon is typical for oligomeric enzyme when monomers are less stable than oligomers.4,5 At the concentration 10 6 mol/L, the rate constants kt and k2 of WT and mutant enzymes were similar. At 1 O 7 mol/L enzyme and lower, the ki and k2 values increased both for the WT and mutant luciferases, but constants ki and k2 of mutant forms were four to six times lower that those of the WT (Table 1). Thus, mutations significantly increased the stability of luciferase at both stages of inactivation. 

Conversion of a-GTP to a-GDP in the simulations occurs with a probability proportional to the a-GTP inactivation rate constant, kh. The inactive a-GDP subunit may combine with a fiy subunit to reform the complete G-protein ... 

The apparent inactivation rate constants of the enzyme, can be estimated from a series of experiments in which each of several concentrations of inhibitor [/] are used. For each [/] the slope of enzyme activity versus preincubation time is measured. The array of kot,s values is then plotted against inhibitor concentrations [/]. The Kiappis the [/] that elicits a ki aa/2. Graphic representations of these experiments are shown in Figure 8.29. 

Boltzmann universal constant first-order inactivation rate constant Planck universal constant catalytic rate constant catalytic rate constant at infinite dilution reaction rate constants according to reaction scheme molar concentration of product molar concentration of competitive inhibitor product molar concentration of non-competitive inhibitor product... 

The inactivation rate constant and its temperature dependence can be experimentally determined as already presented in section 3.5.2. 

Specific activity of biocatalyst molar concentration of substrate B (alternatively coefficient in Eq. 5.3) initial molar concentration of substrate B coefficient in Eq. 5.3 concentration of biocatalyst time of a cycle of reactor operation enzyme activity initial enzyme activity molar concentration of enzyme species Eij volumetric activity of enzyme species Ey enzyme volumetric activity initial enzyme volumetric activity bioreactor feed flow-rate total flow-rate to downstream operations initial feed flow-rate to bioreactor i number of half-lives of biocatalyst use film volumetric mass transfer coefficient for substrate Michaelis-Menten constant catalytic rate constant first-order inactivation rate constant transition rate constants... 

The temperature and pressure dependency of the inactivation rate constants can be described, respectively, by means of Arrhenius equation at constant pressure... 

The inactivation rate constants at reference temperature To and pressure po are given by kr and kj, , respectively. is the activation energy and is the activation volume, both derived from the corresponding slopes of the semilogarithmic plots [2j. Alternatively, for the first-order kinetics, the thermal death time and pressure death time approach can be applied (compare with Eq. (9.8)) with Dr and respectively being the decimal reduction times at reference temperature and pressure [2,46—48]. 

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