Abstract
BackgroundBiological routes for ethylene glycol production have been developed in recent years by constructing the synthesis pathways in different microorganisms. However, no microorganisms have been reported yet to produce ethylene glycol naturally.ResultsXylonic acid utilizing microorganisms were screened from natural environments, and an Enterobacter cloacae strain was isolated. The major metabolites of this strain were ethylene glycol and glycolic acid. However, the metabolites were switched to 2,3-butanediol, acetoin or acetic acid when this strain was cultured with other carbon sources. The metabolic pathway of ethylene glycol synthesis from xylonic acid in this bacterium was identified. Xylonic acid was converted to 2-dehydro-3-deoxy-d-pentonate catalyzed by d-xylonic acid dehydratase. 2-Dehydro-3-deoxy-d-pentonate was converted to form pyruvate and glycolaldehyde, and this reaction was catalyzed by an aldolase. d-Xylonic acid dehydratase and 2-dehydro-3-deoxy-d-pentonate aldolase were encoded by yjhG and yjhH, respectively. The two genes are part of the same operon and are located adjacent on the chromosome. Besides yjhG and yjhH, this operon contains four other genes. However, individually inactivation of these four genes had no effect on either ethylene glycol or glycolic acid production; both formed from glycolaldehyde. YqhD exhibits ethylene glycol dehydrogenase activity in vitro. However, a low level of ethylene glycol was still synthesized by E. cloacae ΔyqhD. Fermentation parameters for ethylene glycol and glycolic acid production by the E. cloacae strain were optimized, and aerobic cultivation at neutral pH were found to be optimal. In fed batch culture, 34 g/L of ethylene glycol and 13 g/L of glycolic acid were produced in 46 h, with a total conversion ratio of 0.99 mol/mol xylonic acid.ConclusionsA novel route of xylose biorefinery via xylonic acid as an intermediate has been established.
Highlights
Biological routes for ethylene glycol production have been developed in recent years by constructing the synthesis pathways in different microorganisms
Screening of xylonic acid utilizing microorganisms Xylonic acid utilizing microorganisms were enriched from soil samples and 4 colonies with different morphologies were isolated from LB agar plates and cultured in flasks
Gene recombination method development Red recombinase assisted gene replacement of E. cloacae was developed as shown in the Method section based on the method we developed in K. pneumoniae [11]. pIJ790 is a plasmid that contains the red recombinase genes and used in E. coli for gene recombination [12]
Summary
Biological routes for ethylene glycol production have been developed in recent years by constructing the synthesis pathways in different microorganisms. Zhang et al Microb Cell Fact (2020) 19:89 dehydratase, 2-dehydro-3-deoxy-d-pentonate aldolase, and aldehyde reductase, respectively. This strain produced 11.7 g/L ethylene glycol from 40 g/L xylose and glycolic acid as a by-product of this process [2]. A synthetic pathway of xylose → xylulose → xylulose-1P → glycolaldehyde → ethylene glycol was constructed in E. coli to produce ethylene glycol from xylose [3] Following these strategies, other pentoses were used as substrates for ethylene glycol synthesis in E. coli [4]. This synthesis pathway was constructed in Corynebacterium glutamicum and E. coli by using serine as an intermediate [5, 6]
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