Efficient recombinant protein production and secretion from nuclear transgenes in Chlamydomonas reinhardtii

Lauersen KJ, Berger H, Mussgnug JH, Kruse O (2013)
Journal of Biotechnology 167(2): 101-110.

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Abstract
Microalgae are diverse photosynthetic microbes which offer the potential for production of a number of high value products (HVP) such as pigments, oils, and bio-active compounds. Fast growth rates, ease of photo-autotrophic cultivation, unique metabolic properties and continuing progress in algal transgenics have raised interest in the use of microalgae systems for recombinant protein (RP) production. This work demonstrates the development of an advanced RP production and secretion system for the green unicellular model alga Chlamydomonas reinhardtii. We generated a versatile expression vector that employs the secretion signal of the native extracellular C. reinhardtii carbonic anhydrase for efficient RP secretion into the culture medium. Unique restriction sites were placed between the regulatory elements to allow fast and easy sub-cloning of sequences of interest. Positive transformants can rapidly be identified by high-throughput plate-level screens via a coupled Gaussia luciferase marker. The vector was tested in Chlamydomonas wild type CC-1883 (WT) and in the transgene expression transformant UVM4. Compared to the native secretion signal of the Gaussia luciferase, up to 84% higher RP production could be achieved. With this new expression system we could generate transformants that express up to 10 mg RP per liter culture without further optimization. The target RP is found exclusively in culture medium and can therefore easily be isolated and purified. We conclude that this new expression system will be a valuable tool for many heterologous protein expression applications from C. reinhardtii in the future.
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Lauersen KJ, Berger H, Mussgnug JH, Kruse O. Efficient recombinant protein production and secretion from nuclear transgenes in Chlamydomonas reinhardtii. Journal of Biotechnology. 2013;167(2):101-110.
Lauersen, K. J., Berger, H., Mussgnug, J. H., & Kruse, O. (2013). Efficient recombinant protein production and secretion from nuclear transgenes in Chlamydomonas reinhardtii. Journal of Biotechnology, 167(2), 101-110.
Lauersen, K. J., Berger, H., Mussgnug, J. H., and Kruse, O. (2013). Efficient recombinant protein production and secretion from nuclear transgenes in Chlamydomonas reinhardtii. Journal of Biotechnology 167, 101-110.
Lauersen, K.J., et al., 2013. Efficient recombinant protein production and secretion from nuclear transgenes in Chlamydomonas reinhardtii. Journal of Biotechnology, 167(2), p 101-110.
K.J. Lauersen, et al., “Efficient recombinant protein production and secretion from nuclear transgenes in Chlamydomonas reinhardtii”, Journal of Biotechnology, vol. 167, 2013, pp. 101-110.
Lauersen, K.J., Berger, H., Mussgnug, J.H., Kruse, O.: Efficient recombinant protein production and secretion from nuclear transgenes in Chlamydomonas reinhardtii. Journal of Biotechnology. 167, 101-110 (2013).
Lauersen, Kyle J., Berger, Hanna, Mussgnug, Jan H., and Kruse, Olaf. “Efficient recombinant protein production and secretion from nuclear transgenes in Chlamydomonas reinhardtii”. Journal of Biotechnology 167.2 (2013): 101-110.
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8 Citations in Europe PMC

Data provided by Europe PubMed Central.

Genetic tools and techniques for Chlamydomonas reinhardtii.
Mussgnug JH., Appl. Microbiol. Biotechnol. 99(13), 2015
PMID: 26025017
Targeted expression of nuclear transgenes in Chlamydomonas reinhardtii with a versatile, modular vector toolkit.
Lauersen KJ, Kruse O, Mussgnug JH., Appl. Microbiol. Biotechnol. 99(8), 2015
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Protein N-glycosylation in eukaryotic microalgae and its impact on the production of nuclear expressed biopharmaceuticals.
Mathieu-Rivet E, Kiefer-Meyer MC, Vanier G, Ovide C, Burel C, Lerouge P, Bardor M., Front Plant Sci 5(), 2014
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Integration of carbon assimilation modes with photosynthetic light capture in the green alga Chlamydomonas reinhardtii.
Berger H, Blifernez-Klassen O, Ballottari M, Bassi R, Wobbe L, Kruse O., Mol Plant 7(10), 2014
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Ice recrystallization inhibition mediated by a nuclear-expressed and -secreted recombinant ice-binding protein in the microalga Chlamydomonas reinhardtii.
Lauersen KJ, Vanderveer TL, Berger H, Kaluza I, Mussgnug JH, Walker VK, Kruse O., Appl. Microbiol. Biotechnol. 97(22), 2013
PMID: 24037309

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