Comparison of Three Xylose Pathways in Pseudomonas putida KT2440 for the Synthesis of Valuable Products.

Isabel Bator,Till Tiso,Andreas Wittgens,Frank Rosenau,Lars M Blank

doi:10.3389/fbioe.2019.00480

Abstract

Pseudomonas putida KT2440 is a well-established chassis in industrial biotechnology. To increase the substrate spectrum, we implemented three alternative xylose utilization pathways, namely the Isomerase, Weimberg, and Dahms pathways. The synthetic operons contain genes from Escherichia coli and Pseudomonas taiwanensis. For isolating the Dahms pathway in P. putida KT2440 two genes (PP_2836 and PP_4283), encoding an endogenous enzyme of the Weimberg pathway and a regulator for glycolaldehyde degradation, were deleted. Before and after adaptive laboratory evolution, these strains were characterized in terms of growth and synthesis of mono-rhamnolipids and pyocyanin. The engineered strain using the Weimberg pathway reached the highest maximal growth rate of 0.30 h−1. After adaptive laboratory evolution the lag phase was reduced significantly. The highest titers of 720 mg L−1 mono-rhamnolipids and 30 mg L−1 pyocyanin were reached by the evolved strain using the Weimberg or an engineered strain using the Isomerase pathway, respectively. The different stoichiometries of the three xylose utilization pathways may allow engineering of tailored chassis for valuable bioproduct synthesis.

Highlights

For the establishment of a circular bioeconomy, the chemical industry has to overcome the massive and ever-increasing use of fossil resources and the concomitant production of environmental pollutions including greenhouse gases
The direct conversion of 2-oxoglutarate into glutamate would benefit from the Weimberg pathway. This was computed by flux balance analysis (FBA) with a maximal product yield of 1 mmol mmol−1 for the Weimberg pathway, and only 0.83 and 0.75 mmol mmol−1 for the Isomerase and the Dahms pathway, respectively
The intermediate glycolaldehyde, using the Dahms pathway for xylose degradation, itself is a relevant chemical for industrial applications and a precursor for ethylene glycol synthesis

Summary

Introduction

For the establishment of a circular bioeconomy, the chemical industry has to overcome the massive and ever-increasing use of fossil resources and the concomitant production of environmental pollutions including greenhouse gases. The alternative is CO2 as carbon source, either directly or fixed via chemocatalysis or plants (Olah et al, 2009; Goeppert et al, 2012). CO2-fixation though cannot only proceed under natural conditions, there are many synthetic approaches to convert CO2 to valuable products. It immediately becomes clear that the knowledge about CO2-fixation pathways can be utilized to redirect the metabolic flow into the production of chemicals using the synthetic biology arsenal. On the one hand cell-free systems are used to fix CO2 via multienzyme cascades (Schwander et al, 2016; Satagopan et al, 2017), on the other hand

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Conclusion