title: Elimination of truncated recombinant protein expressed in Escherichia coli by removing cryptic translation initiation site.
abstract: Undesirable truncated recombinant protein products pose a special expression and purification challenge because such products often share similar chromatographic properties as the desired full length protein. We describe here our observation of both full length and a truncated form of a yeast protein (Gcn5) expressed in Escherichia coli, and the reduction or elimination of the truncated form by mutating a cryptic Shine-Dalgarno or START codon within the Gcn5 coding region. Unsuccessful attempts to engineer in a cryptic translation initiation site into other recombinant proteins suggest that cryptic Shine-Dalgarno or START codon sequences are necessary but not sufficient for cryptic translation in E. coli.
authors.affiliation: Center for Eukaryotic Gene Regulation, Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Jennings
authors.affiliation: Center for Eukaryotic Gene Regulation, Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA.
authors.forename: Adam F
authors.initials: AF
authors.lastname: Barrios
authors.affiliation: Center for Eukaryotic Gene Regulation, Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA. Electronic address: sxt30@psu.edu.
authors.forename: Song
authors.initials: S
authors.lastname: Tan
mesh_terms: D001483:Base Sequence; D018387:Codon, Initiator / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D005786:Gene Expression Regulation; D051548:Histone Acetyltransferases / Q000096:biosynthesis / Q000235:genetics*; D009154:Mutation; D010442:Peptide Chain Initiation, Translational; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000235:genetics*; D000099224:p300-CBP-Associated Factor
chemical_list: D018387:Codon, Initiator; D051548:Histone Acetyltransferases; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C501392:GCN5 protein, S cerevisiae; D000099224:p300-CBP-Associated Factor
keywords: Cryptic initiation; Escherichia coli expression; Recombinant protein expression