Abstract
A Lolium perenne ice-binding protein (LpIBP) demonstrates superior ice recrystallization inhibition (IRI) activity and has proposed applications in cryopreservation, food texturing, as well as in being a “green” gas hydrate inhibitor. Recombinant production of LpIBP has been previously conducted in bacterial and yeast systems for studies of protein characterization, but large-scale applications have been hitherto limited due to high production costs. In this work, a codon-optimized LpIBP was recombinantly expressed and secreted in a novel one-step vector system from the nuclear genome of the green microalga Chlamydomonas reinhardtii. Both mixotrophic and photoautotrophic growth regimes supported LpIBP expression, indicating the feasibility of low-cost production using minimal medium, carbon dioxide, and light energy as input. In addition, multiple growth and bioproduct extraction cycles were performed by repetitive batch cultivation trials, demonstrating the potential for semi-continuous production and biomass harvesting. Concentrations of recombinant protein reached in this proof of concept approach were sufficient to demonstrate IRI activity in culture media without additional purification or concentration, with activity further verified by thermal hysteresis and morphology assays. The incorporation of the recombinant LpIBP into a model gas hydrate offers the promise that algal production may eventually find application as a “green” hydrate inhibitor.
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Acknowledgments
The authors would like to acknowledge the CLIB Graduate Cluster Industrial Biotechnology (Federal Ministry of Science & Technology North Rhine Westphalia, Germany (to K.J.L.)), the Deutsche Forschungsgemeinschaft (DFG KR-1586/5-2 to H.B.) for funding and a Natural Sciences and Engineering Research Council of Canada Discovery grant. The authors would also like to express thanks to Julie Choi and Kristy Moniz for their assistance with ice crystal analyses, and Angelika Schemel and Dr. Sonja Siwiora for assistance with culture media processing. The authors would also like to thank Prof. R. Bock for providing C. reinhardtii strain UVM4.
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Lauersen, K.J., Vanderveer, T.L., Berger, H. et al. Ice recrystallization inhibition mediated by a nuclear-expressed and -secreted recombinant ice-binding protein in the microalga Chlamydomonas reinhardtii . Appl Microbiol Biotechnol 97, 9763–9772 (2013). https://doi.org/10.1007/s00253-013-5226-x
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DOI: https://doi.org/10.1007/s00253-013-5226-x