The Pichia pastoris transmembrane protein GT1 is a glycerol transporter and relieves the repression of glycerol on AOX1 expression

Chunjun Zhan, Songwei Wang, Yang Sun, Xiaofeng Dai, Xiuxia Liu, Linda Harvey, Brian McNeil, Yankun Yang, Zhonghu Bai

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Abstract

Promoter of alcohol oxidase I (PAOX1) is the most efficient promoter involved in the regulation of recombinant protein expression in Pichia pastoris (P. pastoris). PAOX1 is tightly repressed by the presence of glycerol in the culture medium; thus, glycerol must be exhausted before methanol can be taken up by P. pastoris and the expression of the heterologous protein can be induced. In this study, a candidate glycerol transporter (GT1, GeneID: 8197545) was identified, and its role was confirmed by further studies (e.g. bioinformatics analysis, heterologous complementation in Schizosaccharomyces pombe (S. pombe)). When GT1 is co-expressed with enhanced green fluorescent protein (EGFP), it localizes to the membrane and S. pombe carrying gt1 but not the wild-type strain can grow on medium containing glycerol as the sole carbon source. The present study is the first to report that AOX1 in the X-33gt1 mutant can achieve constitutive expression in medium containing glycerol; thus, knocking down gt1 can eliminate the glycerol repression of PAOX1 in P. pastoris. These results suggest that the glycerol transporter may participate in the process of PAOX1 inhibition in glycerol medium.

Original languageEnglish
Pages (from-to)1-10
Number of pages10
JournalFEMS Yeast Research
Volume16
Issue number4
Early online date11 Apr 2016
DOIs
Publication statusPublished - 1 Jun 2016

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Keywords

  • constitutive expression
  • glycerol transporter
  • repression
  • Pichia pastoris
  • release

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