title: The Pichia pastoris transmembrane protein GT1 is a glycerol transporter and relieves the repression of glycerol on AOX1 expression.
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-33Δgt1 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.
authors.affiliation: National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
authors.forename: Chunjun
authors.initials: C
authors.lastname: Zhan
authors.affiliation: National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
authors.forename: Songwei
authors.initials: S
authors.lastname: Wang
authors.affiliation: National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Sun
authors.affiliation: National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
authors.forename: Xiaofeng
authors.initials: X
authors.lastname: Dai
authors.affiliation: National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
authors.forename: Xiuxia
authors.initials: X
authors.lastname: Liu
authors.affiliation: Fermentation Centre, Institute of Pharmacy and Biomedical Science, Strathclyde University, 161 Cathedral Street, Glasgow G1 1XQ, Scotland, UK.
authors.forename: Linda
authors.initials: L
authors.lastname: Harvey
authors.affiliation: Fermentation Centre, Institute of Pharmacy and Biomedical Science, Strathclyde University, 161 Cathedral Street, Glasgow G1 1XQ, Scotland, UK.
authors.forename: Brian
authors.initials: B
authors.lastname: McNeil
authors.affiliation: National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
authors.forename: Yankun
authors.initials: Y
authors.lastname: Yang
authors.affiliation: National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China baizhonghu@jiangnan.edu.cn.
authors.forename: Zhonghu
authors.initials: Z
authors.lastname: Bai
mesh_terms: D000429:Alcohol Oxidoreductases / Q000096:biosynthesis*; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects*; D005990:Glycerol / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D010843:Pichia / Q000235:genetics / Q000378:metabolism*
chemical_list: D026901:Membrane Transport Proteins; D000429:Alcohol Oxidoreductases; C019643:alcohol oxidase; D005990:Glycerol
keywords: PAOX1; Pichia pastoris; constitutive expression; glycerol transporter; release; repression