Evaluation of a new episomal vector based on the GAP promoter for structural genomics in pichia pastoris

In Pyo Hong, Stephen Anderson, Shin Geon Choi

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

A new constitutive episomal expression vector, pGAPZ-E, was constructed and used for initial screening of eukaryotic target gene expression in Pichia pastoris. Two reporter genes such as beta-galactosidase gene and GFPuv gene were overexpressed in P. pastoris. The expression level of the episomal pGAPZ-E strain was higher than that of the integrated form when the beta-galactosidase gene was used as the reporter gene in P. pastoris X33. The avoiding of both the integration procedure and an induction step simplified the overall screening process for eukaryotic target gene expression in P. pastoris. Nine human protein targets from the Core 50, family of Northeast Structural Genomics Consortium (http:// www.nesg.org), which were intractable when expressed in E. coli, were subjected to rapid screening for soluble expression in P. pastoris. HR547, HR919, and HR1697 hurnan proteins, which had previously been found to express poorly or to be insoluble in E. coli, expressed in soluble form in P. pastoris. Therefore, the new episomal GAP promoter vector provides a convenient and alternative system for high-throughput screening of eukaryotic protein expression in P. pastoris.

Original languageEnglish (US)
Pages (from-to)1362-1368
Number of pages7
JournalJournal of Microbiology and Biotechnology
Volume16
Issue number9
StatePublished - Sep 1 2006

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Applied Microbiology and Biotechnology

Keywords

  • Episomal vector
  • Eukaryotic structural genomics
  • GAP promoter
  • Heterologous expression system
  • Pichia pastoris

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