Improving recombinant eukaryotic membrane protein yields in Pichia pastoris: the importance of codon optimization and clone selection

Fredrik Öberg, Jennie Sjöhamn, Matthew T. Conner, Roslyn M. Bill, Kristina Hedfalk

Research output: Contribution to journalArticle

Abstract

In the last 15 years, 80% of all recombinant proteins reported in the literature were produced in the bacterium, Escherichia coli, or the yeast, Pichia pastoris. Nonetheless, developing effective general strategies for producing recombinant eukaryotic membrane proteins in these organisms remains a particular challenge. Using a validated screening procedure together with accurate yield quantitation, we therefore wished to establish the critical steps contributing to high yields of recombinant eukaryotic membrane protein in P. pastoris. Whilst the use of fusion partners to generate chimeric constructs and directed mutagenesis have previously been shown to be effective in bacterial hosts, we conclude that this approach is not transferable to yeast. Rather, codon optimization and the preparation and selection of high-yielding P. pastoris clones are effective strategies for maximizing yields of human aquaporins.
Original languageEnglish
Pages (from-to)398-411
Number of pages14
JournalMolecular Membrane Biology
Volume28
Issue number6
DOIs
Publication statusPublished - Sep 2011

Keywords

  • aquaporins
  • codon
  • electroporation
  • humans
  • lithium chloride
  • pichia
  • recombinant proteins
  • genetic transformation

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