Abstract
Candida tropicalis has recently emerged as a valuable yeast species with respect to lipid metabolism, not only for its oleaginous characteristics but also for its ability to utilize diverse range of substrates. Hence, it can be explored as an ideal host for lipid metabolic engineering, although inadequate genetic transformation system for developing the stable transformant has limited this scope. To resolve this existing constraint, we have come up with a novel strategy of a genomic integrating system in the oleaginous strain SY005 of C. tropicalis. Employing this system, comprising of host-specific regulatable promoter, transcription terminator, and integration locus, we have first examined the expression of a reporter gene, and then ectopically expressed a transcription factor CtRAP1 encoding the repressor activator protein 1 of C. tropicalis SY005. A maximum lipid content of 0.37 g/g dry cell weight was achieved in the engineered strain upon galactose induction, leading to 60% (w/w) increase relative to the wild type strain SY005. This work demonstrates the use of a markerless integrative transformation system to promote lipid accumulation in the diploid yeast without applying nutrient stress and hampering cell growth. The findings of this study will augment the research on lipid metabolic engineering and exploit the enormous potential of C. tropicalis as an industrial lipid feedstock.
Key points •A transformation system was established in oleaginous yeast Candida tropicalis SY005 •Activity of host-specific molecular elements was verified by reporter gene expression •SY005 was engineered to ectopically express a transcriptional regulator gene CtRAP1 •The engineered strain exhibited 60% increase in lipid content on galactose induction •The increase in lipid content was correlated with the induced expression of CtRAP1 |
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Acknowledgments
The authors sincerely thank Dr. Kaustuv Sanyal (Jawaharlal Nehru Centre for Advanced Scientific Research, India) for providing the pURA-del and pBS-CaURA3 plasmids, and Dr. Judith Berman (Department of Microbiology, University of Minnesota, USA) for the pMG2254 plasmid. Authors also acknowledge the grant supports from SGIRG-SRIC (IIT/SRIC/BT/GRT/2014) and Food Security Project (F. No. 4-25/2013-TS-1) of IIT Kharagpur for developing research infrastructure.
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AC: conceptualization, methodology, investigation, formal analysis, validation, writing—original draft. AG: investigation, methodology. MKM: conceptualization, project administration, resources, supervision, writing—review and editing. All authors read and approved the manuscript.
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Chattopadhyay, A., Gupta, A. & Maiti, M.K. Engineering an oleaginous yeast Candida tropicalis SY005 for enhanced lipid production. Appl Microbiol Biotechnol 104, 8399–8411 (2020). https://doi.org/10.1007/s00253-020-10830-6
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DOI: https://doi.org/10.1007/s00253-020-10830-6