Benefits of enzyme kinetics modelling

Mathematical models, especially when coupled with modern computer techniques, prove to be very effective in searching for optimal operating conditions and creating an optimal microenvironment for the biocatalyst in order to optimise productivity. Therefore, the study of the theoretical model for the enzyme reaction system is of interest for the industrial application of the biocatalyst. Theoretical modelling of the enzyme kinetics and the reactors can be used to find optimal operation points and to increase our knowledge about the process. The model usually contains information on the particular biocatalytic reactions. It also includes the mass balance equations in the reactor. The model has to be effective for a wide range of variations of the internal process variables.The benefits of enzyme kinetics modelling are demonstrated in this review with some examples. These are: continuous (R)-mandelic acid production in the enzyme membrane reactor, enzymatic dipeptide synthesis, the synthesis of L-tert-leucine, the synthesis of CMP-Neu5Ac, the synthesis of L-erythrulose, the resolution of racemic alcohol and the synthesis of trisaccharide catalysed by a multi-enzyme system

Modeling of the pyruvate production with Escherichia coli in a fed-batch bioreactor

A family of 10 competing, unstructured models has been developed to model cell growth, substrate consumption, and product formation of the pyruvate producing strain Escherichia coli YYC202 ldhA::Kan strain used in fed-batch processes. The strain is completely blocked in its ability to convert pyruvate into acetyl-CoA or acetate (using glucose as the carbon source) resulting in an acetate auxotrophy during growth in glucose minimal medium. Parameter estimation was carried out using data from fed-batch fermentation performed at constant glucose feed rates of q(VG)=10 mL h(-1). Acetate was fed according to the previously developed feeding strategy. While the model identification was realized by least-square fit, the model discrimination was based on the model selection criterion (MSC). The validation of model parameters was performed applying data from two different fed-batch experiments with glucose feed rate q(VG)=20 and 30 mL h(-1), respectively. Consequently, the most suitable model was identified that reflected the pyruvate and biomass curves adequately by considering a pyruvate inhibited growth (Jerusalimsky approach) and pyruvate inhibited product formation (described by modified Luedeking-Piret/Levenspiel term)

Modelling of L-DOPA enzymatic oxidation catalyzed by L-amino acid oxidases from Crotalus adamanteus and Rhodococcus opacus

L-amino acid oxidases (L-AAO) are well known for their broad substrate specificity. L-amino acid oxidases from Crotalus adamanteus and Rhodococcus opacus were applied for biotransformation of 3,4-dihydroxyphenyl-L-alanine (L-DOPA) as a substrate to its corresponding alpha-keto acid. In this reaction, hydrogen peroxide formed as a by-product causes chemical decarboxylation of alpha-keto acids and acts as competitive product inhibitor. Beef liver catalase was used to decompose it.It was shown that both enzymes were able to oxidize L-DOPA to corresponding products. L-AAO from R. opacus was more specific (lower K I-DOPA value) and more active towards L-DOPA substrate than L-AAO from C. adamanteus. Its catalytic constant, k(3), estimated by Levenspiel's method, was found to be 10-fold higher than the one for L-AAO from C adamanteus. L-AAO from R. opacus exhibits slightly L-DOPA inhibition, which is not the case for L-AAO from C adamanteus.The biotransformations Of L-DOPA were carried out in batch enzyme membrane reactor (EMR), as well as in the repetitive batch EMR. The reactor and kinetics were modelled. Parameters were estimated by differential and integral method and presented in this article. (c) 2005 Elsevier B.V. All rights reserved

Process strategies to enhance pyruvate production with recombinant Escherichia coli: From repetitive fed-batch to in situ product recovery with fully integrated electrodialysis

Using the pyruvate production strain Escherichia coli YYC202 IdhA::Kan different process alternatives are studied with the aim of preventing potential product inhibition by appropriate product separation. This strain is completely blocked in its ability to convert pyruvate into acetyl-CoA or acetate, resulting in acetate auxotrophy during growth in glucose minimal medium. Continuous experiments with cell retention, repetitive fed-batch, and an in situ product recovery (ISPR) process with fully integrated electrodialysis were tested. Although the continuous approach achieved a high volumetric productivity (Q(P)) of 110 g L-1 d(-1), this approach was not pursued because of long-term production strain instabilities. The highest pyruvate/glucose molar yield of up to 1.78 mol mol(-1) together with high Q(P) 145 g L-1 d(-1) and high pyruvate titers was achieved by the repetitive fed-batch approach. To separate pyruvate from fermentation broth a fully integrated continuous process was developed. In this process electrodialysis was used as a separation unit. Under optimum conditions a (calculated) final pyruvate titer of >900 mmol L-1 (79 g L-1) was achieved. (C) 2004 Wiley Periodicals, Inc