Production of d -Glyceric acid from d -Galacturonate in Escherichia coli
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METABOLIC ENGINEERING AND SYNTHETIC BIOLOGY - ORIGINAL PAPER
Production of d‑Glyceric acid from d‑Galacturonate in Escherichia coli Kevin J. Fox1 · Kristala L. J. Prather1 Received: 24 May 2020 / Accepted: 7 October 2020 © Society for Industrial Microbiology and Biotechnology 2020
Abstract A microbial production platform has been developed in Escherichia coli to synthesize d-glyceric acid from d-galacturonate. The expression of uronate dehydrogenase (udh) from Pseudomonas syringae and galactarolactone isomerase (gli) from Agrobacterium fabrum, along with the inactivation of garK, encoding for glycerate kinase, enables d-glyceric acid accumulation by utilizing the endogenous expression of galactarate dehydratase (garD), 5-keto-4-deoxy-D-glucarate aldolase (garL), and 2-hydroxy-3-oxopropionate reductase (garR). Optimization of carbon flux through the elimination of competing metabolic pathways led to the development of a ΔgarKΔhyiΔglxKΔuxaC mutant strain that produced 4.8 g/l of d-glyceric acid from d-galacturonate, with an 83% molar yield. Cultivation in a minimal medium produced similar yields and demonstrated that galactose or glycerol serve as possible carbon co-feeds for industrial production. This novel platform represents an alternative for the production of d-glyceric acid, an industrially relevant chemical, that addresses current challenges in using acetic acid bacteria for its synthesis: increasing yield, enantio-purity and biological stability. Keywords Metabolic engineering · Escherichia coli · d-glyceric acid
Introduction Production of commodity chemicals is currently highly dependent on fossil fuels, a non-renewable and environmentally detrimental feedstock [1]. Finding alternative, renewable starting materials to synthesize these products allows for a more secure production outlook. Renewable resources bring new challenges to chemical production, however, including logistical challenges due to their often de-centralized production as well as the increased complexity of the material itself [2]. Food waste, which is rich in diverse carbon sources and is at a surplus in many parts of the world, is seen as a promising renewable feedstock [3–5]. Food wastes contain a diverse set of sugars naturally structured into pectin, hemicellulose or cellulose that are valuable resources after chemical or enzymatic pretreatment [6, 7]. Bioprocessing, specifically using metabolically engineered Electronic supplementary material The online version of this article (https://doi.org/10.1007/s10295-020-02323-2) contains supplementary material, which is available to authorized users. * Kristala L. J. Prather [email protected] 1
Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
microbes such as Escherichia coli, is an attractive method to create value-added products from these diverse sugar feeds [8, 9]. Using E. coli as a host organism is attractive due to the relatively high degree of understanding of many of its cellular processes as well as the many natural sugar c
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