Orthologous promoters from related methylotrophic yeasts surpass expression of endogenous promoters of Pichia pastoris

Thomas Vogl, Jasmin Elgin Fischer, Patrick Hyden, Richard Wasmayer, Lukas Sturmberger, Anton Glieder*

*Corresponding author for this work

Research output: Contribution to journalArticle

Abstract

Methylotrophic yeasts such as Komagataella phaffii (syn. Pichia pastoris, Pp), Hansenula polymorpha (Hp), Candida boidinii (Cb) and Pichia methanolica (Pm) are widely used protein production platforms. Typically, strong, tightly regulated promoters of genes coding for their methanol utilization (MUT) pathways are used to drive heterologous gene expression. Despite highly similar open reading frames in the MUT pathways of the four yeasts, the regulation of the respective promoters varies strongly between species. While most endogenous Pp MUT promoters remain tightly repressed after depletion of a repressing carbon, Hp, Cb and Pm MUT promoters are derepressed to up to 70% of methanol induced levels, enabling methanol free production processes in their respective host background. Here, we have tested a series of orthologous promoters from Hp, Cb and Pm in Pp. Unexpectedly, when induced with methanol, the promoter of the HpMOX gene reached very similar expression levels as the strong methanol, inducible, and most frequently used promoter of the Pp alcohol oxidase 1 gene (PPpAOX1). The HpFMD promoter even surpassed PPpAOX1 up to three-fold, when induced with methanol, and reached under methanol-free/derepressed conditions similar expression as the methanol induced PPpAOX1. These results demonstrate that orthologous promoters from related yeast species can give access to otherwise unobtainable regulatory profiles and may even considerably surpass endogenous promoters in P. pastoris.

Original languageEnglish
Article number38
JournalAMB Express
Volume10
Issue number1
DOIs
Publication statusPublished - 1 Dec 2020

Keywords

  • Derepression
  • Komagataella phaffii
  • Methanol-free
  • Orthologous promoters
  • Pichia pastoris

ASJC Scopus subject areas

  • Biophysics
  • Applied Microbiology and Biotechnology

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