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Improvement of sabinene tolerance of Escherichia coli using adaptive laboratory evolution and omics technologies
- Source :
- Biotechnology for Biofuels, Vol 13, Iss 1, Pp 1-15 (2020)
- Publication Year :
- 2020
- Publisher :
- Springer Science and Business Media LLC, 2020.
-
Abstract
- Background Biosynthesis of sabinene, a bicyclic monoterpene, has been accomplished in engineered microorganisms by introducing heterologous pathways and using renewable sugar as a carbon source. However, the efficiency and titers of this method are limited by the low host tolerance to sabinene (in both eukaryotes and prokaryotes). Results In this study, Escherichia coli BL21(DE3) was selected as the strain for adaptive laboratory evolution. The strain was evolved by serial passaging in the medium supplemented with gradually increasing concentration of sabinene, and the evolved strain XYF(DE3), which exhibited significant tolerance to sabinene, was obtained. Then, XYF(DE3) was used as the host for sabinene production and an 8.43-fold higher sabinene production was achieved compared with the parental BL21(DE3), reaching 191.76 mg/L. Whole genomes resequencing suggested the XYF(DE3) strain is a hypermutator. A comparative analysis of transcriptomes of XYF(DE3) and BL21(DE3) was carried out to reveal the mechanism underlying the improvement of sabinene tolerance, and 734 up-regulated genes and 857 down-regulated genes were identified. We further tested the roles of the identified genes in sabinene tolerance via reverse engineering. The results demonstrated that overexpressions of ybcK gene of the DLP12 family, the inner membrane protein gene ygiZ, and the methylmalonyl-CoA mutase gene scpA could increase sabinene tolerance of BL21(DE3) by 127.7%, 71.1%, and 75.4%, respectively. Furthermore, scanning electron microscopy was applied to monitor cell morphology. Under sabinene stress, the parental BL21(DE3) showed increased cell length, whereas XYF(DE3) showed normal cell morphology. In addition, overexpression of ybcK, ygiZ or scpA could partially rescue cell morphology under sabinene stress and overexpression of ygiZ or scpA could increase sabinene production in BL21(DE3). Conclusions This study not only obtained a sabinene-tolerant strain for microbial production of sabinene but also revealed potential regulatory mechanisms that are important for sabinene tolerance. In addition, for the first time, ybcK, ygiZ, and scpA were identified to be important for terpene tolerance in E. coli BL21(DE3).
- Subjects :
- 0106 biological sciences
lcsh:Biotechnology
Monoterpene
Heterologous
Sabinene
Management, Monitoring, Policy and Law
Biology
medicine.disease_cause
Cell morphology
01 natural sciences
Applied Microbiology and Biotechnology
lcsh:Fuel
Sabinene tolerance
03 medical and health sciences
chemistry.chemical_compound
Mutase
lcsh:TP315-360
lcsh:TP248.13-248.65
010608 biotechnology
Escherichia coli
medicine
Transcriptome sequencing
Gene
030304 developmental biology
0303 health sciences
ybcK/ygiZ/scpA
Strain (chemistry)
Renewable Energy, Sustainability and the Environment
Genome resequencing
General Energy
chemistry
Biochemistry
bacteria
Adaptive laboratory evolution
Biotechnology
Subjects
Details
- ISSN :
- 17546834
- Volume :
- 13
- Database :
- OpenAIRE
- Journal :
- Biotechnology for Biofuels
- Accession number :
- edsair.doi.dedup.....3e3ba489928672d2c8c341c4b8c14419