Expression of the Kirsten ras viral and human proteins in Escherichia coli.
The expression vectors pINIII-A and pINIII (lpp p5) were used to construct plasmids which direct the synthesis in Escherichia coli of the Kirsten ras viral (v-Ki-ras) and human cellular (c-Ki-ras) oncogene products as fusion proteins containing 9 and 10 extra amino acids, respectively, at their N termini. Authenticity of the bacterially produced proteins was determined by immunoprecipitation and immunoblot analyses with ras-specific monoclonal antibodies. After induction with isopropyl-beta-D-thiogalactopyranoside, the viral protein represented approximately 20% of the total cellular protein. The majority of the protein was found in the postsonication low-speed centrifugation pellet. The synthesized viral protein was active in GTP binding, as judged by autophosphorylation and photoaffinity labeling assays.
D004262:DNA Restriction Enzymes; D004926:Escherichia coli / Q000235:genetics*; D005796:Genes; D005822:Genetic Vectors; D006801:Humans; D007708:Kirsten murine sarcoma virus / Q000235:genetics*; D015689:Oncogene Protein p21(ras); D009856:Oncogene Proteins, Viral / Q000235:genetics*; D009857:Oncogenes; D010957:Plasmids; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras); D009053:Sarcoma Viruses, Murine / Q000235:genetics*; D014764:Viral Proteins / Q000235:genetics*
D009856:Oncogene Proteins, Viral; D011518:Proto-Oncogene Proteins; D014764:Viral Proteins; D004262:DNA Restriction Enzymes; C501009:HRAS protein, human; D015689:Oncogene Protein p21(ras); D016283:Proto-Oncogene Proteins p21(ras)