Engineered Fusion Proteins for Efficient Protein Secretion and Purification of a Human Growth Factor from the Green Microalga Chlamydomonas reinhardtii

Thomas Baier, Dana Kros, Rebecca C. Feiner, Kyle J. Lauersen, Kristian M. Müller, Olaf Kruse

Research output: Contribution to journalArticlepeer-review

24 Scopus citations

Abstract

Light-driven recombinant protein (RP) production in eukaryotic microalgae offers a sustainable alternative to other established cell-culture systems. RP production via secretion into the culture medium enables simple product separation from the cells adding a layer of process value in addition to the algal biomass, which can be separately harvested. For the model microalga Chlamydomonas reinhardtii, a broad range of molecular tools have been established to enable heterologous gene expression; however, low RP production levels and unreliable purification from secretion concepts have been reported. Domesticated C. reinhardtii strains used for genetic engineering are often cell-wall deficient. These strains nevertheless secrete cell-wall components such as insoluble (hydroxy)proline-rich glycoproteins into the culture media, which hinder downstream purification processes. Here, we attempted to overcome limitations in secretion titers and improve protein purification by combining fusion partners that enhance RP secretion and enable alternative aqueous two-phase (ATPS) RP extraction from the culture medium. Protein fusions were strategically designed to contain a stably folded peptide, which enhanced secretion capacities and gave insights into (some) regulatory mechanisms responsible for this process in the algal host. The elevated protein titers mediated by this fusion were then successfully applied in combination with a fungal hydrophobin tag, which enabled protein purification from the complex microalgal extracellular environment by ATPS, to yield functional recombinant human epidermal growth factor (hEGF) from the algal host.
Original languageEnglish (US)
JournalACS Synthetic Biology
Volume7
Issue number11
DOIs
StatePublished - Nov 16 2018
Externally publishedYes

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