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Engineering Pseudomonas putida KT2440 for the production of isobutanol

Abstract : We engineered P. putida for the production of isobutanol from glucose by preventing product and precursor degradation, inactivation of the soluble transhydrogenase SthA, overexpression of the native ilvC and ilvD genes, and implementation of the feedbackresistant acetolactate synthase AlsS from Bacillus subtilis, ketoacid decarboxylase KivD from Lactococcus lactis, and aldehyde dehydrogenase YqhD from Escherichia coli. The resulting strain P. putida Iso2 produced isobutanol with a substrate specific product yield (Y Iso/S) of 22 ± 2 mg per gram of glucose under aerobic conditions. Furthermore, we identified the ketoacid decarboxylase from Carnobacterium maltaromaticum to be a suitable alternative for isobutanol production, since replacement of kivD from L. lactis in P. putida Iso2 by the variant from C. maltaromaticum yielded an identical Y Iso/S. Although P. putida is regarded as obligate aerobic, we show that under oxygen deprivation conditions this bacterium does not grow, remains metabolically active, and that engineered producer strains secreted isobutanol also under the non-growing conditions.
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Submitted on : Friday, April 16, 2021 - 10:48:16 AM
Last modification on : Thursday, January 6, 2022 - 11:38:05 AM
Long-term archiving on: : Saturday, July 17, 2021 - 6:26:40 PM


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Robert Nitschel, Andreas Ankenbauer, Ilona Welsch, Nicolas Wirth, Christoph Massner, et al.. Engineering Pseudomonas putida KT2440 for the production of isobutanol. Engineering in Life Sciences, Wiley-VCH Verlag, 2020, 20, pp.148 - 159. ⟨10.1002/elsc.201900151⟩. ⟨hal-02529614⟩



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