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Model-based optimisation of the cascade synthesis of L-homoserine

One-pot cascade synthesis of L-homoserine catalysed by transaminase and pyruvate-dependent aldolase starting from pyruvate, formaldehyde and L-alanine (Scheme 1) was investigated in this work. The proof of concept was shown to work before by Hernandez et al. (2017). In this work two types of biocatalysts were used and kinetically characterized ; i.e. separate enzymes available as cell free extract (CFE). The focus was put on kinetic investigation of this complex reaction. Detailed kinetic analysis was done and kinetic parameters were estimated from the experimental data collected by using the initial reaction rate method. Based on the reaction scheme and the estimated apparent kinetic parameters the kinetic model was developed and was combined with mass balances in the reactor for separate reaction steps, as well as for the cascade reaction. Scheme 1 The reaction scheme of the synthesis of L-homoserine. The proposed model described the data well for the separate reaction steps ; i.e. aldol addition of pyruvate and formaldehyde catalyzed by aldolase and transamination of L-alanine by the product of the aldol addition. The kinetic model of the cascade reaction was used to find and propose the strategy for the improvement of product concentration and volume productivity. It was found that the best way to perform the reaction is in the fed-batch reactor with continuous feed of formaldehyde and two additions of L-alanine and pyruvate during the course of experiment up to their initial concentrations. This strategy enabled the production of L- homoserine of 672 mM and 76.23 g L-1 d-1 after 25 hours. The kinetic modeling was found to be crucial for understanding of the reaction system, and the obtained process metrics present a good starting point for potential industrial application. K. Hernandez, J. Bujons, J. Joglar, S. J. Charnock, P. Domínguez de María, W. Dieter Fessner, P. Clapés, ACS Catal., 2017, 7(3), 1707- 1711, doi: https://doi.org/10.1021/acscatal.6b03181. Project CARBAZYMES-This project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 635595.

cascade reaction, mathematical modeling, process optimization

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