Novel molecular biological tools for the efficient expression of fungal lytic polysaccharide monooxygenases in Pichia pastoris

Lukas Rieder, Katharina Ebner, Anton Glieder, Morten Sørlie*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

Background: Lytic polysaccharide monooxygenases (LPMOs) are attracting large attention due their ability to degrade recalcitrant polysaccharides in biomass conversion and to perform powerful redox chemistry. Results: We have established a universal Pichia pastoris platform for the expression of fungal LPMOs using state-of-the-art recombination cloning and modern molecular biological tools to achieve high yields from shake-flask cultivation and simple tag-less single-step purification. Yields are very favorable with up to 42 mg per liter medium for four different LPMOs spanning three different families. Moreover, we report for the first time of a yeast-originating signal peptide from the dolichyl-diphosphooligosaccharide-protein glycosyltransferase subunit 1 (OST1) form S. cerevisiae efficiently secreting and successfully processes the N-terminus of LPMOs yielding in fully functional enzymes. Conclusion: The work demonstrates that the industrially most relevant expression host P. pastoris can be used to express fungal LPMOs from different families in high yields and inherent purity. The presented protocols are standardized and require little equipment with an additional advantage with short cultivation periods.

Original languageEnglish
Article number122
JournalBiotechnology for Biofuels
Volume14
Issue number1
DOIs
Publication statusPublished - Dec 2021

Keywords

  • LPMO
  • Pichia pastoris
  • Signal peptide cleaving
  • Simplified expression

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology
  • Renewable Energy, Sustainability and the Environment
  • Energy(all)
  • Management, Monitoring, Policy and Law

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