Ectoine Production Using Novel Heterologous EctABCS. salarius from Marine Bacterium Salinicola salarius
Source abstract
Abstract
Ectoine, a heterocyclic amino acid produced by various bacteria, was widely used in the fields of cosmetics and medicine. In this study, a novel ectoine synthesis cluster from marine bacterium Salinicola salarius 1A01339 was firstly introduced into Escherichia coli BL21(DE3) for heterologous production of ectoine. The bioinformatic analysis proved the function of these ectoine synthesis enzymes, and showed the highest identities of 83.3–87.7% with enzymes from other microorganisms. Using the whole-cell biocatalytic method, 3.28 g/L ectoine was synthesized and excreted into the medium with the substrate of 200 mM sodium aspartate at 25 °C, pH 6.5 in flask-level. Further bioconversion was performed in the fermentor system at the high cell density of 20 OD/mL, and the concentration of extracellular ectoine was increased to 22.5 g/L in 24 h (equivalent to the specific productivity of 0.94 g/L·h), achieving over 6 times of production compared with that in flasks. Significantly, the recombinant strain demonstrated a lower catalytic temperature with the optimum of 25 °C, and a stronger tolerance to the substrate aspartate of 300 mM. These results might provide a compelling case for ectoine synthesis as well as potential applications in large-scale industrial production.
Source: crossref · Quality: baseline
Research context
OpenAlex topics: Microbial Metabolic Engineering and Bioproduction · Enzyme Catalysis and Immobilization · Enzyme Structure and Function
SciVX discovery tags are terminology-based. OpenAlex topics are source-provided classifications.
Affiliations
Anhui Provincial Key Laboratory of Molecular Enzymology and Mechanism of Major Diseases, 1 Beijing East Road, Wuhu 241000, China
Key Laboratory of Biomedicine in Gene Diseases and Health of Anhui Higher Education Institutes, College of Life Sciences, Anhui Normal University, 1 Beijing East Road, Wuhu 241000, China
Funding
University Natural Science Research Project of Anhui Province
References (43)
Show source-provided references
1. Intracellular ion and organic solute concentrations of the extremely halophilic bacterium Salinibacter ruber10.1007/s00792-002-0286-3 ↗
2. Fenizia, S., Thume, K., Wirgenings, M., and Pohnert, G. (2020). Ectoine from Bacterial and Algal Origin is a Compatible Solute in Microalgae. Mar. Drugs, 18.10.3390/md18010042 ↗
3. 1,4,5,6-Tetrahydro-2-methyl-4-pyrimidinecarboxylic acid. A novel cyclic amino acid from halophilic phototrophic bacteria of the genus Ectothiorhodospira10.1111/j.1432-1033.1985.tb08903.x ↗
4. Characterization of biosynthetic enzymes for ectoine as a compatible solute in a moderately halophilic eubacterium, Halomonas elongata10.1128/JB.181.1.91-99.1999 ↗
5. Characterization of genes for the biosynthesis of the compatible solute ectoine from Marinococcus halophilus and osmoregulated expression in Escherichia coli10.1099/00221287-143-4-1141 ↗
6. Stressed out: Bacterial response to high salinity using compatible solute biosynthesis and uptake systems, lessons from Vibrionaceae10.1016/j.csbj.2021.01.030 ↗
7. Metabolic Pathway Construction and Optimization of Escherichia coli for High-Level Ectoine Production10.1007/s00284-020-01888-6 ↗
8. Nasal spray and eye drops containing ectoine, a novel natural, non-drug anti-allergic substance are not less effective than azelastine nasal spray and eye drops in improving the symptoms of allergic rhinitis and conjunctivitis
9. Microbial production of ectoine and hydroxyectoine as high-value chemicals10.1186/s12934-021-01567-6 ↗
10. The Biosynthesis of Ectoine10.1111/j.1574-6968.1990.tb03815.x ↗
11. Osmotically induced synthesis of the compatible solute hydroxyectoine is mediated by an evolutionarily conserved ectoine hydroxylase10.1074/jbc.M704023200 ↗
12. Rational flux-tuning of Halomonas bluephagenesis for co-production of bioplastic PHB and ectoine10.1038/s41467-020-17223-3 ↗
13. Current status on metabolic engineering for the production of l-aspartate family amino acids and derivatives10.1016/j.biortech.2017.05.145 ↗
14. The ups and downs of ectoine: Structural enzymology of a major microbial stress protectant and versatile nutrient10.1515/hsz-2020-0223 ↗
15. Degradation of the microbial stress protectants and chemical chaperones ectoine and hydroxyectoine by a bacterial hydrolase-deacetylase complex10.1074/jbc.RA120.012722 ↗
16. Heterologous Ectoine Production in Escherichia coli: Optimization Using Response Surface Methodology10.1155/2019/5475361 ↗
17. Industrial Production of the Cell Protectant Ectoine: Protection Mechanisms, Processes, and Products10.2174/22115501113026660037 ↗
18. Contribution of mechanosensitive channels to osmoadaptation and ectoine excretion in Halomonas elongata10.1007/s00792-020-01168-y ↗
19. High production of ectoine from aspartate and glycerol by use of whole-cell biocatalysis in recombinant Escherichia coli10.1186/s12934-015-0238-0 ↗
20. New type of osmoregulated solute transporter identified in halophilic members of the Bacteria domain: TRAP transporter TeaABC mediates uptake of ectoine and hydroxyectoine in Halomonas elongata DSM 2581(T)10.1128/JB.184.11.3078-3085.2002 ↗
21. A blueprint of ectoine metabolism from the genome of the industrial producer Halomonas elongata DSM 2581(T)10.1111/j.1462-2920.2010.02336.x ↗
22. Bacterial milking: A novel bioprocess for production of compatible solutes10.1002/(SICI)1097-0290(19980205)57:3<306::AID-BIT7>3.0.CO;2-L ↗
23. Identification and characterization of ectoine biosynthesis genes and heterologous expression of the ectABC gene cluster from Halomonas sp. QHL1, a moderately halophilic bacterium isolated from Qinghai Lake10.1007/s12275-014-3389-5 ↗
24. Metabolic Engineering of Corynebacterium glutamicum for High-Level Ectoine Production: Design, Combinatorial Assembly, and Implementation of a Transcriptionally Balanced Heterologous Ectoine Pathway10.1002/biot.201800417 ↗
25. A Novel Cold-adapted Endoglucanase (M6A) from Microbacterium kitamiense S12 Isolated from Qinghai-Tibetan Plateau10.1007/s12257-019-0009-6 ↗
26. MEGA11: Molecular Evolutionary Genetics Analysis Version 1110.1093/molbev/msab120 ↗
27. The architecture of the diaminobutyrate acetyltransferase active site provides mechanistic insight into the biosynthesis of the chemical chaperone ectoine10.1074/jbc.RA119.011277 ↗
28. The crystal structure of the tetrameric DABA-aminotransferase EctB, a rate-limiting enzyme in the ectoine biosynthesis pathway10.1111/febs.15265 ↗
29. Illuminating the catalytic core of ectoine synthase through structural and biochemical analysis10.1038/s41598-018-36247-w ↗
30. Pathway construction and metabolic engineering for fermentative production of ectoine in Escherichia coli10.1016/j.ymben.2016.02.013 ↗
31. Clustal W and clustal X version 2.010.1093/bioinformatics/btm404 ↗
32. Deciphering key features in protein structures with the new ENDscript server10.1093/nar/gku316 ↗
33. GCN5-related histone N-acetyltransferases belong to a diverse superfamily that includes the yeast SPT10 protein10.1016/S0968-0004(97)01034-7 ↗
34. Crystal structure of human recombinant ornithine aminotransferase10.1006/jmbi.1997.1583 ↗
35. Osmotically regulated synthesis of the compatible solute ectoine in Bacillus pasteurii and related Bacillus spp.10.1128/AEM.68.2.772-783.2002 ↗
36. Czech, L., Hermann, L., Stoveken, N., Richter, A.A., Hoppner, A., Smits, S.H.J., Heider, J., and Bremer, E. (2018). Role of the Extremolytes Ectoine and Hydroxyectoine as Stress Protectants and Nutrients: Genetics, Phylogenomics, Biochemistry, and Structural Analysis. Genes, 9.10.3390/genes9040177 ↗
37. Continuous synthesis and excretion of the compatible solute ectoine by a transgenic, nonhalophilic bacterium10.1128/AEM.02482-06 ↗
38. Margesin, R. (2017). Psychrophiles: From Biodiversity to Biotechnology, Springer.10.1007/978-3-319-57057-0 ↗
39. Molecular cloning, expression, and biochemical characterization of a novel cold-active alpha-amylase from Bacillus sp. dsh19-110.1007/s00792-018-1034-7 ↗
40. Highly cold-active pectinases under wine-like conditions from non-Saccharomyces yeasts for enzymatic production during winemaking10.1111/lam.12390 ↗
41. Boosting Escherichia coli’s heterologous production rate of ectoines by exploiting the non-halophilic gene cluster from Acidiphilium cryptum10.1007/s00792-020-01188-8 ↗
42. Exploiting Substrate Promiscuity of Ectoine Hydroxylase for Regio- and Stereoselective Modification of Homoectoine10.3389/fmicb.2019.02745 ↗
43. Biosynthesis of the Stress-Protectant and Chemical Chaperon Ectoine: Biochemistry of the Transaminase EctB10.3389/fmicb.2019.02811 ↗