Skip to main content
SpringerLink
Log in

Identification of a Long-Chain Fatty Acid Elongase from Nannochloropsis sp. Involved in the Biosynthesis of Fatty Acids by Heterologous Expression in Saccharomyces cerevisiae

  • Published:
Journal of Ocean University of China Aims and scope Submit manuscript

Abstract

The marine microalga Nannochloropsis sp. contains various elongases and desaturases that are critical for biosynthesis of polyunsaturated fatty acids. A full-length cDNA encoding a long-chain fatty acid elongase, named NsFAE, was cloned from Nannochloropsis sp.. The open reading frame of NsFAE (GenBank accession no. MF680548) consisted of 1068bp and encoded a predicted protein of 355 amino acids with molecular mass 38.8kDa. The deduced polypeptide showed 43%-44% identity to fatty acyl elongases from other algae. RT-PCR experiments indicated that the NsFAE gene exhibited the highest expression in Nannochloropsis sp. at 72h (i.e., during the third growth stage) and the expression was significantly lower in the other four growth stages. Plasmid pNsFAE-CRISPR and a recombinant DNA fragment (ADH1p-NsFAE-CYCt) were transformed into Saccharomyces cerevisiae strain BY4742 using the CRISPR-Cas system. Yeast transformants containing NsFAE produced three fatty acids not normally present in wild-type BY4742-linoleic acid, linolenic acid and eicosadienoic acid-indicating that NsFAE encodes a functional elongase enzyme.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Aslan M. A., and Ilmutdin M. A., 2017. The study of microal-gae Nannochloropsis salina fatty acid composition of the extracts using different techniques. SCF vs conventional extraction. Journal of Molecular Liquids, 239: 96–100, DOI: https://doi.org/10.1016/j.molliq.

    Article  Google Scholar 

  • Bao Z., Xiao H., Liang J., Zhang L., Xiong X., Sun N., Si T., and Zhao H., 2015. A Homology Integrated CRISPR-Cas (HI-CRISPR) system for one-step multi-gene disruptions in Saccharomyces cerevisiae. ACS Synthetic Biology, 4: 585–594, DOI: https://doi.org/10.1021/sb500255k.

    Article  Google Scholar 

  • Baoxiu Q., Fraser T., Mugford S., Dobson G., Sayanova O., Butler J., Napier J. A., Stobart A. K., and Lazarus C. M., 2004. Production of very long chain polyunsaturated omega-3 and omega-6 fatty acids in plants. Nature Biotechnology, 22: 739–745, DOI: https://doi.org/10.1038/nbt972.

    Article  Google Scholar 

  • Chodok, P., Eiamsaard, P., Cove D. J., Quatrano R. S., and Kaewsuwan S., 2013. Identification and functional characterization of two Δ12-fatty acid desaturases associated with essential linoleic acid biosynthesis in Physcomitrella patens. Journal of Industrial Microbiology and Biotechnology, 40: 901–913, DOI: https://doi.org/10.1007/s10295-013-1285-3.

    Article  Google Scholar 

  • Christopher L., Brian, P., Ruben F. M., and Nancy A. D. S., 2015. Overproduction and secretion of free fatty acids through disrupted neutral lipid recycle in Saccharomyces cerevisiae. Metabolic Engineering, 28: 54–62, DOI: https://doi.org/10.1016/j.ymben.2014.11.006.

    Article  Google Scholar 

  • Corteggiani C. E., Telatin A., Vitulo N., Forcato C., D'Angelo M., Schiavon R., Vezzi A., Giacometti G. M., Morosinotto T., and Valle G., 2014. Chromosome scale genome assembly and transcriptome profiling of Nannochloropsis gaditana in nitrogen depletion. Molecular Plant, 7: 323–335, DOI: https://doi.org/10.1093/mp/sst120.

    Article  Google Scholar 

  • Dong H. K., Periasamy A., Young S. J., Bidur P. C., Jeong W. S., and Byung K. H., 2010. Identification and characterization of a novel enzyme related to the synthesis of PUFAs derived from Thraustochytrium aureum ATCC 34304. Biotechnology and Bioprocess Engineering, 15: 261–272, DOI: https://doi.org/10.1007/s12257-009-0223-8.

    Article  Google Scholar 

  • Faten B. A., Mohamed B., Fatma E., Nesrine K., Mouna D., Bruno B., Pierre, V., Slim A., and Imen F., 2017. Cyanobacteria as source of marine bioactive compounds: Molecular specific detection based on Δ9 desaturase gene. International Journal of Biological Macromolecules, 105: 1440–1445.

    Article  Google Scholar 

  • Guan B., Lei J., Su S., Chen F., Duan Z., Chen, Y., Gong X., Li H., and Jin J., 2012. Absence of Yps7p, a putative glycosylphosphatidylinositol-linked aspartyl protease in Pichia pastoris, results in aberrant cell wall composition and increased osmotic stress resistance. FEMS Yeast Research, 12: 969–979, DOI:https://doi.org/10.1111/1567-1364.12002.

    Article  Google Scholar 

  • Jun X., Ling W., and Jian-bo, Z., 2016. Expression of Shewanella frigidimarina fatty acid metabolic genes in E. coli by CRISPR/cas9-coupled lambda red recombineering. Biotechnology Letters, 38: 117–122, DOI: https://doi.org/10.1007/s10529-015-1956-4.

    Article  Google Scholar 

  • Junichiro O., Keishi S., Yuji O., Nozomu O., and Makoto I., 2013. Two fatty acid elongases possessing C18-Δ6/C18-Δ9/C20-Δ5 or C16-Δ9 elongase activity in Thraustochytrium sp. ATCC 26185. Marine Biotechnology, 15: 476–486, DOI: https://doi.org/10.1007/s10126-013-9496-1.

    Article  Google Scholar 

  • Kendall A. C., Kiezel T. M., Brownbridge L. C., Harwood J. L., and Nicolaou A., 2017. Lipid functions in skin: Differential effects of n-3 polyunsaturated fatty acids on cutaneous ceramides, in a human skin organ culture model. BBA-Biomembranes, 1859: 1679–1689, DOI: https://doi.org/10.1016/j.bbamem.2017.03.016.

    Article  Google Scholar 

  • Kaewsuwan S., Bunyapraphatsara N., Cove D. J., Quatrano R. S., and Chodok P., 2010. High level production of adrenic acid in Physcomitrella patens using the algae Pavlova sp. Delta(5)-elongase gene. Bioresource Technology, 101: 4081–4088, DOI: https://doi.org/10.1016/j.biortech.2009.12.138.

    Article  Google Scholar 

  • Leonard A. E., Pereira S. L., and Sprecher H., 2004. Elongation of long-chain fatty acids. Progress in Lipid Research, 43: 36–54, DOI: https://doi.org/10.1016/S0163-7827(03)00040-7.

    Article  Google Scholar 

  • Leonie W., Tao Y., and Florian D., 2017. Establishing very long-chain fatty alcohol and wax ester biosynthesis in Saccharomyces cerevisiae. Biotechnology and Bioengineering, 114: 1025–1035, DOI: https://doi.org/10.1002/bit.26220.

    Article  Google Scholar 

  • Li M., Ou X. Y., Yang X. D., Guo D. Q., Qian X. Y., Xing L. J., and Li M. C., 2012. Cloning and identification of a novel C18-Δ9 polyunsaturated fatty acid specific elongase gene from DHA-producing Isochrysis galbana H29. Biotechnology and Bioprocess Engineering, 17: 22–32, DOI:https://doi.org/10.1007/s12257-011-0037-3.

    Article  Google Scholar 

  • Lieselot B., Charlotte B., Koen, G., Céline, D., and Imogen F., 2018. Influence of high pressure homogenization on free fatty acid formation in Nannochloropsis sp. European Journal of Lipid Science and Technology, 120: 1–19, DOI: https://doi.org/10.1002/ejlt.201700436.

    Google Scholar 

  • Lina J. D., Camille R., Giorgio, P., Guillaume T., Richard B., Marina L., Tomas M., Frédérique, T., Jean D. F., Denis F., Juliette J., Olga S., Frédéric, B., and Eric M., 2017. A palmitic acid elongase affects eicosapentaenoic acid and plastidial monogalactosyldiacylglycerol levels in Nannochloropsis. Plant Physiology, 173: 742–759, DOI: https://doi.org/10.1104/pp.16.01420.

    Article  Google Scholar 

  • Montes R., Chisaguano A. M., and Castellote A. I., 2013. Fatty-acid composition of maternal and umbilical cord plasma and early childhood atopic eczema in a Spanish cohort. European Journal of Clinical Nutrition, 67: 658–663, DOI: https://doi.org/10.1038/ejcn.2013.68.

    Article  Google Scholar 

  • Niu, Y., Kong J., Fu L., Yang J., and Xu Y., 2009. Identification of a novel C20-elongase gene from the marine microalgae Pavlova viridis and its expression in Escherichia coli. Mar Biotechnol, 11: 17–23, DOI: https://doi.org/10.1007/s10126-008-9116-7.

    Article  Google Scholar 

  • Nicolás, L., Sebastian M., María, P. C., Natalia R., Dante T., Alex D. G., Natalia, G., Nicole E., and Alejandro M., 2017. Reconstruction of the microalga Nannochloropsis salina genome-scale metabolic model with applications to lipid production. BMC Systems Biology, 11: 2–17, DOI: https://doi.org/10.1186/s12918-017-0441-1.

    Google Scholar 

  • Norashikin N., Loh S. H., Aziz A., and Cha T. S., 2018. Metabolic engineering of fatty acid biosynthesis in Chlorella vulgaris using an endogenous omega-3 fatty acid desaturase gene with its promoter. Algal Research, 31: 262–275, DOI: https://doi.org/10.1016/j.algal.2018.02.020.

    Article  Google Scholar 

  • Petrie J. R., Mackenzie A. M., Shrestha P., Liu Q., Frampton D. F., Robert S. S., and Singh S. P., 2010. Isolation of three novel long-chain polyunsaturated fatty acid delta 9-elongases and the transgenic assembly of the entire Pavlova salina docosahexaenoic and pathway in Nicotiana benthamianal. Journal of Phycology, 46: 917–925, DOI: https://doi.org/10.1111/j.1529-8817.2010.00870.x.

    Article  Google Scholar 

  • Phatthanon P., Yoshinobu K., Minetaka S., Takeshi B., Eii-chiro, F., Akio K., and Satoshi H., 2007. Functional analysis of very long-chain fatty acid elongase gene, HpELO2, in the methylotrophic yeast Hansenula polymorpha. Applied Microbiology and Biotechnology, 76: 417–427, DOI: https://doi.org/10.1007/s00253-007-1012-y.

    Article  Google Scholar 

  • Qi B., Beaudoin F., Fraser T., Stobart A. K., Napier J. A., and Lazarus C. M., 2002. Identification of a cDNA encoding a novel C18-Δ9 polyunsaturated fatty acid-specific elongating activity from the docosahexaenoic acid (DHA)-producing microalga, Isochrysis galbana. FEBS Letters, 510: 159–162, DOI: https://doi.org/10.1016/S0014-5793(01)03247-1.

    Article  Google Scholar 

  • Rebecca M. L., and Brian F. P., 2012. Engineering Escherichia coli to synthesize free fatty acids. Trends in Biotechnology, 30: 659–667, DOI: https://doi.org/10.1016/j.tibtech.2012.09.006.

    Article  Google Scholar 

  • Robert S. S., Singh S. P., and Rong Z. X., 2005. Metabolic engineering of Arabidopsis to produce nutritionally important DHA in seed oil. Functional Plant Biology, 32: 473–479, DOI: https://doi.org/10.1071/FP05084.

    Article  Google Scholar 

  • Sakuradani E., Nojiri M., Suzuki H., and Shimizu S., 2009. Identification of a novel fatty acid elongase with a wide substrate specificity from arachidonic acid-producing fungus Mortierella alpina 1S-4. Applied Microbiology and Biotechnology, 84: 709–716, DOI: https://doi.org/10.1007/s00253-009-1999-3.

    Article  Google Scholar 

  • Shi T. L., Yu, A. Q., Li M., Ou X. Y., Xing L. J., and Li M. C., 2012. Identification of a novel C22-Delta 4-producing docosahexaenoic acid (DHA) specific polyunsaturated fatty acid desaturase gene from Isochrysis galbana and its expression in Saccharomyces cerevisiae. Biotechnology Letters, 34: 2265–2274, DOI: https://doi.org/10.1007/s10529-012-1028-y.

    Article  Google Scholar 

  • Shi T. L., Yu A. Q., Li M., Zhang M., Xing L. J., and Li M. C., 2013. Identification and characterization of a novel C20-elongase gene from the marine microalgae, Pavlova viridis, and its use for the reconstitution of two pathways of longchain polyunsatured fatty acids biosynthesis in Saccharomyces cerevisiae. Biotechnology Letters, 35: 1271–1282, DOI: https://doi.org/10.1007/s10529-013-1194-6.

    Article  Google Scholar 

  • Shi H. B., Wu M., Zhu J. J., Zhang C. H., Yao D. W., Luo J., and Loor J. J., 2017. Fatty acid elongase 6 plays a role in the synthesis of long-chain fatty acids in goat mammary epithelial cells. Journal of Dairy Science, 100: 4987–4995, DOI: https://doi.org/10.3168/jds.2016-12159.

    Article  Google Scholar 

  • Sushil B., Joon N. L., and Young-Il K., 2016. The fatty acid chain elongase, Elovl1, is required for kidney and swim bladder development during zebrafish embryogenesis. Organogenesis, 12: 78–93, DOI: https://doi.org/10.1080/15476278.2016.1172164.

    Article  Google Scholar 

  • Sun H. K., So Y. K., Eui K. K., Kyung H. R., Jung B. K., and Kwang S. K., 2014. Identification and functional characterization of polyunsaturated fatty acid elongase (McELOVL5) gene from pike eel (Muraenesox cinereus). Biotechnology Letters, 36: 29–37, DOI: https://doi.org/10.1007/s10529-013-1344-x.

    Article  Google Scholar 

  • Takamiya M., Sakurai M., and Teranishi F., 2016. Lead discovery for mammalian elongation of long chain fatty acids family 6 using a combination of high-throughput fluorescent-based assay and RapidFire mass spectrometry assay. Biochemical and Biophysical Research Communications, 480: 721–726, DOI: https://doi.org/10.1016/j.bbrc.2016.10.103.

    Article  Google Scholar 

  • Takeuchi T., 2001. A review of feed development for early life stages of marine finfish in Japan. Aquaculture, 200: 203–222, DOI: https://doi.org/10.1016/S0044-8486(01)00701-3.

    Article  Google Scholar 

  • Xie D. Z., Chen F., Lin S. Y., You C. H., Wang S. Q., Zhang Q. H., Monroig, Ő., Tocher D. R., and Li Y. Y., 2016. Longchain polyunsaturated fatty acid biosynthesis in the euryhaline herbivorous teleost Scatophagus argus: Functional characterization, tissue expression and nutritional regulation of two fatty acyl elongases. Comparative Biochemistry & Physiology Part B, 198: 37–45, DOI: https://doi.org/10.1016/j.cbpb.2016.03.009.

    Article  Google Scholar 

  • Zheng M., Tian J., Yang, G., Zheng L., Chen, G., Chen J., and Wang B., 2013. Transcriptome sequencing, annotation and expression analysis of Nannochloropsis sp. at different growth phases. Gene, 523: 117–121, DOI: https://doi.org/10.1016/j.gene..2013.04.005.

    Article  Google Scholar 

  • Zhu B. H., Tu C. C., Shi H. P., Yang G. P., and Pan K. H., 2017. Overexpression of endogenous delta-6 fatty acid desaturase gene enhances eicosapentaenoic acid accumulation in Phaeodactylum tricornutum. Process Biochemistry, 57: 43–49.

    Article  Google Scholar 

Download references

Acknowledgments

This work was supported by the Basic Scientific Fund for National Public Research Institutes of China (No. 2016Q07).

Author information

Authors and Affiliations

  1. College of Food, Shihezi University, Shihezi, 832000, China

    Minrui Guo, Guogang Chen & Jiluan Chen

  2. The First Institute of Oceanography, State Oceanic Administration of China, Qingdao, 266061, China

    Minggang Zheng

Authors
  1. Minrui Guo
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Guogang Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jiluan Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Minggang Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Guogang Chen.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Guo, M., Chen, G., Chen, J. et al. Identification of a Long-Chain Fatty Acid Elongase from Nannochloropsis sp. Involved in the Biosynthesis of Fatty Acids by Heterologous Expression in Saccharomyces cerevisiae. J. Ocean Univ. China 18, 1199–1206 (2019). https://doi.org/10.1007/s11802-019-3946-y

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11802-019-3946-y

Key words

Search

Navigation