title: Ubiquitin fusion augments the yield of cloned gene products in Escherichia coli.
abstract: Despite the availability of efficient transcription and translation signals, some heterologous gene products are not adequately expressed when introduced into prokaryotes and eukaryotes. An expression system has been established in Escherichia coli to increase the yield of cloned gene products, where the C terminus of ubiquitin was fused to the N terminus of unstable or poorly expressed proteins. Fusion of ubiquitin to yeast metallothionein or to the alpha subunit of the adenylate cyclase-stimulatory GTP-binding protein increased the yield from undetectable to 20% of the total cellular protein. A ubiquitin-N alpha-protein hydrolase has been partially purified from rabbit reticulocytes; this enzyme faithfully cleaves the junction peptide bound between the C-terminal Gly-76 of ubiquitin and the fusion protein. The increased yield of cloned gene products is very likely due to increased stability and/or more efficient translation of the fusion proteins. Possible mechanisms for the augmentation of ubiquitin fusion-protein expression in prokaryotes and eukaryotes are discussed.
authors.affiliation: Smith Kline & French Laboratories, Department of Molecular Pharmacology, King of Prussia, PA 19406.
authors.forename: T R
authors.initials: TR
authors.lastname: Butt
authors.forename: S
authors.initials: S
authors.lastname: Jonnalagadda
authors.forename: B P
authors.initials: BP
authors.lastname: Monia
authors.forename: E J
authors.initials: EJ
authors.lastname: Sternberg
authors.forename: J A
authors.initials: JA
authors.lastname: Marsh
authors.forename: J M
authors.initials: JM
authors.lastname: Stadel
authors.forename: D J
authors.initials: DJ
authors.lastname: Ecker
authors.forename: S T
authors.initials: ST
authors.lastname: Crooke
mesh_terms: D003001:Cloning, Molecular; D010450:Endopeptidases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D019204:GTP-Binding Proteins / Q000235:genetics; D005786:Gene Expression Regulation; D008668:Metallothionein / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000502:physiology*; D011994:Recombinant Proteins / Q000502:physiology*; D014452:Ubiquitins / Q000502:physiology*
chemical_list: D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014452:Ubiquitins; D008668:Metallothionein; D010450:Endopeptidases; C059237:ubiquitin-Nalpha-protein hydrolase; D019204:GTP-Binding Proteins