title: Investigating the dynamics of recombinant protein secretion from a microalgal host.
abstract: Production of recombinant proteins with microalgae represents an alternative platform over plant- or bacterial-based expression systems for certain target proteins. Secretion of recombinant proteins allows accumulation of the target product physically separate from the valuable algal biomass. To date, there has been little investigation into the dynamics of recombinant protein secretion from microalgal hosts-the culture parameters that encourage secreted product accumulation and stability, while encouraging biomass production. In this work, the efficiency of recombinant protein production was optimized by adjusting cultivation parameters for a strain of Chlamydomonas reinhardtii previously engineered to secrete a functional recombinant Lolium perenne ice binding protein (LpIBP), which has applications as a frozen food texturing and cryopreservation additive, into its culture medium. Three media and several cultivation styles were investigated for effects on secreted LpIBP titres and culture growth. A combination of acetate and carbon dioxide feeding with illumination resulted in the highest overall biomass and recombinant protein titres up to 10mgL(-1) in the culture medium. Pure photoautotrophic production was possible using two media types, with recombinant protein accumulation in all cultivations correlating to culture cell density. Two different cultivation systems were used for scale-up to 10L cultivations, one of which produced yields of secreted recombinant protein up to 12mgL(-1) within six cultivation days. Functional ice recrystallization inhibition (IRI) of the LpIBP from total concentrated extracellular protein extracts was demonstrated in a sucrose solution used as a simplified ice cream model. IRI lasted up to 7 days, demonstrating the potential of secreted products from microalgae for use as food additives.
authors.affiliation: Bielefeld University, Faculty of Biology, Center for Biotechnology (CeBiTec), Universitätsstrasse 27, 33615 Bielefeld, Germany.
authors.forename: Kyle J
authors.initials: KJ
authors.lastname: Lauersen
authors.affiliation: Bielefeld University, Faculty of Biology, Center for Biotechnology (CeBiTec), Universitätsstrasse 27, 33615 Bielefeld, Germany.
authors.forename: Isabel
authors.initials: I
authors.lastname: Huber
authors.affiliation: Bielefeld University, Faculty of Biology, Center for Biotechnology (CeBiTec), Universitätsstrasse 27, 33615 Bielefeld, Germany.
authors.forename: Julian
authors.initials: J
authors.lastname: Wichmann
authors.affiliation: Bielefeld University, Faculty of Biology, Center for Biotechnology (CeBiTec), Universitätsstrasse 27, 33615 Bielefeld, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Baier
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Sciences, Section I: Food Process Engineering, Kaiserstraße 12, 76131 Karlsruhe, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Leiter
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Sciences, Section I: Food Process Engineering, Kaiserstraße 12, 76131 Karlsruhe, Germany.
authors.forename: Volker
authors.initials: V
authors.lastname: Gaukel
authors.affiliation: Institute of Biotechnological Process Engineering, Faculty of Engineering and Mathematics, University of Applied Sciences, Universitätsstrasse 27, 33615 Bielefeld, Germany.
authors.forename: Viktor
authors.initials: V
authors.lastname: Kartushin
authors.affiliation: Institute of Biotechnological Process Engineering, Faculty of Engineering and Mathematics, University of Applied Sciences, Universitätsstrasse 27, 33615 Bielefeld, Germany.
authors.forename: Anke
authors.initials: A
authors.lastname: Rattenholl
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Sciences, Section III: Bioprocess Engineering, Fritz-Haber-Weg 2, 76131 Karlsruhe, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Steinweg
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Sciences, Section III: Bioprocess Engineering, Fritz-Haber-Weg 2, 76131 Karlsruhe, Germany.
authors.forename: Lena
authors.initials: L
authors.lastname: von Riesen
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Sciences, Section III: Bioprocess Engineering, Fritz-Haber-Weg 2, 76131 Karlsruhe, Germany.
authors.forename: Clemens
authors.initials: C
authors.lastname: Posten
authors.affiliation: Institute of Biotechnological Process Engineering, Faculty of Engineering and Mathematics, University of Applied Sciences, Universitätsstrasse 27, 33615 Bielefeld, Germany.
authors.forename: Frank
authors.initials: F
authors.lastname: Gudermann
authors.affiliation: Institute of Biotechnological Process Engineering, Faculty of Engineering and Mathematics, University of Applied Sciences, Universitätsstrasse 27, 33615 Bielefeld, Germany.
authors.forename: Dirk
authors.initials: D
authors.lastname: Lütkemeyer
authors.affiliation: Bielefeld University, Faculty of Biology, Center for Biotechnology (CeBiTec), Universitätsstrasse 27, 33615 Bielefeld, Germany.
authors.forename: Jan H
authors.initials: JH
authors.lastname: Mussgnug
authors.affiliation: Bielefeld University, Faculty of Biology, Center for Biotechnology (CeBiTec), Universitätsstrasse 27, 33615 Bielefeld, Germany. Electronic address: olaf.kruse@uni-bielefeld.de.
authors.forename: Olaf
authors.initials: O
authors.lastname: Kruse
mesh_terms: D018533:Biomass; D002245:Carbon Dioxide; D018929:Cell Culture Techniques; D016825:Chlamydomonas reinhardtii / Q000254:growth & development / Q000378:metabolism; D052836:Heterotrophic Processes; D008029:Lighting; D058086:Microalgae / Q000378:metabolism*; D058433:Photobioreactors; D011994:Recombinant Proteins / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D002245:Carbon Dioxide
keywords: Chlamydomonas reinhardtii; Flat-panel photobioreactor; Lolium perenne ice-binding protein; Microalgae; Recombinant protein secretion; Wave-bag culture