Abstract
Glycolic acid (GA) is an ⍺-hydroxy acid used in cosmetics, packaging, and medical industries due to its excellent properties, especially in its polymeric form. In this study, Escherichia coli was engineered to produce GA from D-xylose by linking the Dahms pathway, the glyoxylate bypass, and the partial reverse glyoxylate pathway (RGP). Initially, a GA-producing strain was constructed by disrupting the xylAB and glcD genes in the E. coli genome and overexpressing the xdh(Cc) from Caulobacter crescentus. This strain was further improved through modular optimization of the Dahms pathway and the glyoxylate bypass. Results for module 1 showed that the rate-limiting step of the Dahms pathway was the xylonate dehydratase reaction, and the overexpression of yagF was sufficient to overcome this bottleneck. Furthermore, the appropriate aldolase gene for module 1 was proven to be yagE. The results also show that overexpression of the lactaldehyde dehydrogenase gene, aldA, is needed to increase the GA production while the overexpression of glyoxylate reductase gene, ycdW, was only essential when the glyoxylate bypass was active. On the other hand, the module 2 enzymes AceA and AceK were vital in activating the glyoxylate bypass, while the RGP enzymes were dispensable. The final strain (GA19) produced 4.57 g/L GA with a yield of 0.46 g/g from D-xylose. So far, this is the highest value achieved for GA production in engineered E. coli through the Dahms pathway.
Similar content being viewed by others
References
Alkim C, Trichez D, Cam Y, Spina L, François JM, Walther T (2016) The synthetic xylulose-1 phosphate pathway increases production of glycolic acid from xylose-rich sugar mixtures. Biotechnol Biofuels 9(1):201. https://doi.org/10.1186/s13068-016-0610-2
Babilas P, Knie U, Abels C (2012) Cosmetic and dermatologic use of alpha hydroxy acids. J German Soc Derm 10(7):488–491. https://doi.org/10.1111/j.1610-0387.2012.07939.x
Baldoma L, Aguilar J (1988) Metabolism of L-fucose and L-rhamnose in Escherichia coli: aerobic-anaerobic regulation of L-lactaldehyde dissimilation. J Bacteriol 170(1):416–421. https://doi.org/10.1128/jb.170.1.416-421.1988
Bhaskar V, Kumar M, Manicka S, Tripathi S, Venkatraman A, Krishnaswamy S (2011) Identification of biochemical and putative biological role of a xenolog from Escherichia coli using structural analysis. Proteins 79(4):1132–1142. https://doi.org/10.1002/prot.22949
Borthwick AC, Holms WH, Nimmo HG (1984) The phosphorylation of Escherichia coli isocitrate dehydrogenase in intact cells. Biochem J 222(3):797–804. https://doi.org/10.1042/bj2220797
Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72(1-2):248–254. https://doi.org/10.1016/0003-2697(76)90527-3
Cabulong RB, Valdehuesa KNG, Ramos KRM, Nisola GM, Lee W-K, Lee CR, Chung W-J (2017) Enhanced yield of ethylene glycol production from D-xylose by pathway optimization in Escherichia coli. Enzym Microb Technol 97:11–20. https://doi.org/10.1016/j.enzmictec.2016.10.020
Cam Y, Alkim C, Trichez D, Trebosc V, Vax A, Bartolo F, Besse P, François JM, Walther T (2016) Engineering of a synthetic metabolic pathway for the assimilation of (D)-xylose into value-added chemicals. ACS Synth Biol 5(7):607–618. https://doi.org/10.1021/acssynbio.5b00103
Cao Y, Niu W, Guo J, Xian M, Liu H (2015) Biotechnological production of 1,2,4-butanetriol: an efficient process to synthesize energetic material precursor from renewable biomass. Sci Rep 5(1):18149. https://doi.org/10.1038/srep18149
Cherepanov PP, Wackernagel W (1995) Gene disruption in Escherichia coli: TcR and KmR cassettes with the option of Flp-catalyzed excision of the antibiotic-resistance determinant. Gene 158(1):9–14. https://doi.org/10.1016/0378-1119(95)00193-A
Choi SY, Kim WJ, Yu SJ, Park SJ, Im SG, Lee SY (2017) Engineering the xylose-catabolizing Dahms pathway for production of poly(d-lactate-co-glycolate) and poly(d-lactate-co-glycolate-co-d-2-hydroxybutyrate) in Escherichia coli. Microb Biotechnol 10(6):29–1364. https://doi.org/10.1111/1751-7915.12721
Choi SY, Park SJ, Kim WJ, Yang JE, Lee H, Shin J, Lee SY (2016) One-step fermentative production of poly(lactate-co-glycolate) from carbohydrates in Escherichia coli. Nat Biotechnol 34(4):435–440. https://doi.org/10.1038/nbt.3485
Chung CT, Niemela SL, Miller RH (1989) One-step preparation of competent Escherichia coli: transformation and storage of bacterial cells in the same solution. Proc Natl Acad Sci U S A 86(7):2172–2175. https://doi.org/10.1073/pnas.86.7.2172
Clark DP, Cronan JE (2013) Two-carbon compounds and fatty acids as carbon sources. https://doi.org/10.1128/ecosalplus.3.4.4
Cronan JE Jr, Laporte D (2013) Tricarboxylic acid cycle and glyoxylate bypass. EcoSal Plus 1(2):1–24. https://doi.org/10.1128/ecosalplus.3.5.2
Dahms AS, Donald A (1982) 2-Keto-3-deoxy-d-xylonate aldolase (3-deoxy-d-pentulosonic acid aldolase). Meth Enzymol 90(Pt E):269–272
Deng Y, Mao Y, Zhang X (2015) Metabolic engineering of E. coli for efficient production of glycolic acid from glucose. Biochem Eng J 103:256–262. https://doi.org/10.1016/j.bej.2015.08.008
Dischert W, Colomb C, Soucaille P (2012) Fermentation process for producing glycolic acid
Fredenberg S, Wahlgren M, Reslow M, Axelsson A (2011) The mechanisms of drug release in poly(lactic-co-glycolic acid)-based drug delivery systems—a review. Int J Pharm 415(1-2):34–52. https://doi.org/10.1016/j.ijpharm.2011.05.049
Green MR, Sambrook J (2012) Molecular cloning: a laboratory manual, 4th edn. Cold Spring Harbor Laboratory Press, New York
Hinago H, Nagahara H, Aoki T (2015) Method for producing glycolic acid
Jiang Y, Liu W, Cheng T, Cao Y, Zhang R, Xian M (2015) Characterization of D-xylonate dehydratase YjhG from Escherichia coli. Bioengineering 6(4):227–232. https://doi.org/10.1080/21655979.2015.1040208
Koivistoinen OM, Kuivanen J, Barth D, Turkia H, Pitkänen J-P, Penttilä M, Richard P (2013) Glycolic acid production in the engineered yeasts Saccharomyces cerevisiae and Kluyveromyces lactis. Microb Cell Factories 12(1):82. https://doi.org/10.1186/1475-2859-12-82
Kornberg HL (1966) The role and control of the glyoxylate cycle in Escherichia coli. Biochem J 99(1):1–11. https://doi.org/10.1042/bj0990001
Limón A, Hidalgo E, Aguilar J (1997) The aldA gene of Escherichia coli is under the control of at least three transcriptional regulators. Microbiol 143(6):2085–2095. https://doi.org/10.1099/00221287-143-6-2085
Liu H, Lu T (2015) Autonomous production of 1,4-butanediol via a de novo biosynthesis pathway in engineered Escherichia coli. Metab Eng 29:135–141. https://doi.org/10.1016/j.ymben.2015.03.009
Liu H, Ramos KRM, Valdehuesa KNG, Nisola GM, Lee W-K, Chung W-J (2013) Biosynthesis of ethylene glycol in Escherichia coli. Appl Microbiol Biotechnol 97(8):3409–3417. https://doi.org/10.1007/s00253-012-4618-7
Liu H, Valdehuesa KNG, Nisola GM, Ramos KRM, Chung W-J (2012) High yield production of D-xylonic acid from D-xylose using engineered Escherichia coli. Bioresour Technol 115:244–248. https://doi.org/10.1016/j.biortech.2011.08.065
Mainguet SE, Gronenberg LS, Wong SS, Liao JC (2013) A reverse glyoxylate shunt to build a non-native route from C4 to C2 in Escherichia coli. Metab Eng 19:116–127. https://doi.org/10.1016/j.ymben.2013.06.004
Meijnen J-P, de Winde JH, Ruijssenaars HJ (2009) Establishment of oxidative D-xylose metabolism in Pseudomonas putida S12. Appl Environ Microbiol 75(9):2784–2791. https://doi.org/10.1128/AEM.02713-08
Nuñez MF, Pellicer MT, Badia J, Aguilar J, Baldoma L (2001) Biochemical characterization of the 2-ketoacid reductases encoded by ycdW and yiaE genes in Escherichia coli. Biochem J 354(3):707–715. https://doi.org/10.1042/bj3540707
Pereira B, Li Z-J, De Mey M, Lim CG, Zhang H, Hoeltgen C, Stephanopoulos G (2016) Efficient utilization of pentoses for bioproduction of the renewable two-carbon compounds ethylene glycol and glycolate. Metab Eng 34:80–87. https://doi.org/10.1016/j.ymben.2015.12.004
Stephens C, Christen B, Fuchs T, Sundaram V, Watanabe K, Jenal U (2007) Genetic analysis of a novel pathway for D-xylose metabolism in Caulobacter crescentus. J Bacteriol 189(5):2181–2185. https://doi.org/10.1128/JB.01438-06
Sun L, Yang F, Sun H, Zhu T, Li X, Li Y, Xu Z, Zhang Y (2016) Synthetic pathway optimization for improved 1,2,4-butanetriol production. J Ind Microbiol Biotechnol 43(1):67–78. https://doi.org/10.1007/s10295-015-1693-7
Tai Y-S, Xiong M, Jambunathan P, Wang J, Wang J, Stapleton C, Zhang K (2016) Engineering nonphosphorylative metabolism to generate lignocellulose-derived products. Nat Chem Biol 12(4):1–10. https://doi.org/10.1038/nchembio.2020
Valdehuesa KNG, Lee W-K, Ramos KRM, Cabulong RB, Choi J, Liu H, Nisola GM, Chung W-J (2015) Identification of aldehyde reductase catalyzing the terminal step for conversion of xylose to butanetriol in engineered Escherichia coli. Bioprocess Biosyst Eng 38(9):1761–1772. https://doi.org/10.1007/s00449-015-1417-4
Valdehuesa KNG, Liu H, Ramos KRM, Park SJ, Nisola GM, Lee W-K, Chung W-J (2014) Direct bioconversion of D-xylose to 1,2,4-butanetriol in an engineered Escherichia coli. Process Biochem 49(1):25–32. https://doi.org/10.1016/j.procbio.2013.10.002
Walsh K, Koshland DE (1984) Determination of flux through the branch point of two metabolic cycles. J Biol Chem 259(15):9646–9654
Wang J, Shen X, Jain R, Wang J, Yuan Q, Yan Y (2017) Establishing a novel biosynthetic pathway for the production of 3,4-dihydroxybutyric acid from xylose in Escherichia coli. Metab Eng 41:39–45. https://doi.org/10.1016/j.ymben.2017.03.003
Xu P, Vansiri A, Bhan N, Koffas MAG (2012) ePathBrick: a synthetic biology platform for engineering metabolic pathways in E. coli. ACS Synth Biol 1(7):256–266. https://doi.org/10.1021/sb300016b
Zahoor A, Otten A, Wendisch VF (2014) Metabolic engineering of Corynebacterium glutamicum for glycolate production. J Biotechnol 192(Pt B):366–375. https://doi.org/10.1016/j.jbiotec.2013.12.020
Zhang N, Wang J, Zhang Y, Gao H (2016) Metabolic pathway optimization for biosynthesis of 1,2,4-butanetriol from xylose by engineered Escherichia coli. Enzym Microb Technol 93-94:51–58. https://doi.org/10.1016/j.enzmictec.2016.07.007
Funding
This work was supported by Korea Research Fellowship Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT (2015H1D3A1062172 and 2016R1C1B1013252) and by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (No. 2009-0093816).
Author information
Authors and Affiliations
Corresponding authors
Ethics declarations
Conflict of interest
The authors declare that they have no conflict of interest.
Ethical approval
This article does not contain any studies with human participants performed by any of the authors.
Electronic supplementary material
ESM 1
(PDF 190 kb)
Rights and permissions
About this article
Cite this article
Cabulong, R.B., Lee, WK., Bañares, A.B. et al. Engineering Escherichia coli for glycolic acid production from D-xylose through the Dahms pathway and glyoxylate bypass. Appl Microbiol Biotechnol 102, 2179–2189 (2018). https://doi.org/10.1007/s00253-018-8744-8
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00253-018-8744-8