title: pCold-GST vector: a novel cold-shock vector containing GST tag for soluble protein production.
abstract: The production of recombinant protein in Escherichia coli is often hampered by low expression levels and low solubility. A variety of methodologies have been developed including protein production at low temperature, and fusion protein expression using soluble protein tags. Here, we present the novel cold-shock vector pCold-GST for high-level expression of soluble proteins in E. coli. This vector is a modified pCold I cold-shock vector that includes the glutathione S-transferase (GST) tag. The pCold-GST expression system developed was applied to 10 proteins that could not be expressed using conventional E. coli expression methodologies, and nine of these proteins were successfully obtained in the soluble fraction. The expression and purification of two unstable protein fragments were also demonstrated by employing a C-terminal hexa-histidine tag for purification purposes. The purified proteins were amenable to NMR analyses. These data suggest that the pCold-GST expression system can be utilized to improve the expression and purification of various proteins.
authors.affiliation: Laboratory of Biophysics, Graduate School of Biological Sciences, Nara Institute of Science and Technology (NAIST), Ikoma, Nara 630 0192, Japan.
authors.forename: Kokoro
authors.initials: K
authors.lastname: Hayashi
authors.forename: Chojiro
authors.initials: C
authors.lastname: Kojima
mesh_terms: D003001:Cloning, Molecular; D005822:Genetic Vectors / Q000737:chemistry* / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics* / Q000378:metabolism; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D015202:Protein Engineering; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D012995:Solubility
chemical_list: D011506:Proteins; D011993:Recombinant Fusion Proteins; D006639:Histidine; D005982:Glutathione Transferase