Accelerated biocatalyst stability testing for process optimization.
The deactivation of protein biocatalysts even at relatively low temperatures is one of the principal drawbacks to their use. To aid in the development of novel biocatalysts, we have derived an equation for both time- and temperature-dependent activity of the biocatalyst based on known concepts such as transition state theory and the Lumry-Eyring model. We then derived an analytical solution for the total turnover number (ttn), under isothermal operation, as a function of the catalytic constant kcat, the unfolding equilibrium constant K, and the intrinsic first-order deactivation rate constant(s) k(d,i). Employing an immobilized glucose isomerase biocatalyst in a CSTR and utilizing a linear temperature ramp beyond the Tm of the enzyme, we demonstrate an accelerated method for extracting the thermodynamic and kinetic constants describing the biocatalyst system. In addition, we demonstrate that the predicted biocatalyst behavior at different temperatures and reaction times is consistent with the experimental observations.
Gibbs
Phillip R
PR
School of Chemical and Biomolecular Engineering, Parker H. Petit Institute of Bioengineering and Biosciences, Georgia Institute of Technology, 315 Ferst Drive, Atlanta, Georgia 30332-0363, USA.
Uehara
Christian S
CS
Neunert
Urban
U
Bommarius
Andreas S
AS
D019747:Aldose-Ketose Isomerases / Q000737:chemistry*; D002384:Catalysis; D003198:Computer Simulation; D004789:Enzyme Activation; D004795:Enzyme Stability; D005632:Fructose / Q000138:chemical synthesis*; D005947:Glucose / Q000737:chemistry*; D007700:Kinetics; D008956:Models, Chemical
D005632:Fructose; D019747:Aldose-Ketose Isomerases; C023298:xylose isomerase; D005947:Glucose