Production of a phosphorylated GST::HPV-6 E7 fusion protein using a yeast expression vector and glutathione S-transferase fusions.
A Saccharomyces cerevisiae GAL7 expression vector for the production of protein fusions to glutathione S-transferase (GST) has been constructed. Using this vector, a GST fusion to human papillomavirus type 6 (HPV-6) E7 protein was produced and purified by affinity chromatography in a single step, at a yield of 2 micrograms/ml of culture. The E7 portion of the fusion protein was phosphorylated, in contrast to the same product made in Escherichia coli. Therefore, yeast GST vectors may be of specific use in producing phosphoproteins, or proteins with other eukaryotic post-translational modifications, in preparative amounts for in vitro analysis.
Romanos
M A
MA
Wellcome Research Laboratories, Beckenham, Kent, UK.
Hughes
F J
FJ
Comerford
S A
SA
Scorer
C A
CA
D005822:Genetic Vectors / Q000235:genetics*; D005982:Glutathione Transferase / Q000096:biosynthesis* / Q000235:genetics; D009856:Oncogene Proteins, Viral / Q000096:biosynthesis* / Q000235:genetics; D027383:Papillomaviridae / Q000235:genetics*; D010750:Phosphoproteins / Q000096:biosynthesis* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D009856:Oncogene Proteins, Viral; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; C062495:oncogene protein E7, Human papillomavirus type 6; D005982:Glutathione Transferase