High-level expression in Escherichia coli of enzymatically active harvey murine sarcoma virus p21ras protein / Expression à haut niveau chez E.c. de la protéine p21ras du virus sarcome murin harvey enzymatiquement active

Le gène de la protéine p21ras est mis sous contrôle du promoteur de transcription du gène cII du bactériophage λ. Synthèse chez E.c. d'une protéine chimère de 23000 daltons contenant tous les résidus d'aminoacides de la protéine p21ras à l'exception des 4 résidus de l'extrémité aminoterminale. Caractéristiques immunologiques et enzymatiques identiques à la protéine virale.

The gene for the Harvey murine sarcoma virus (Ha-MuSV) p21ras protein was fused to the amino-terminal portion of the bacteriophage λ cII gene on the expression vector pJL6. The fusion was such that transcription was controlled by the well-regulated phage λ pL promoter, and translation initiated in the cII gene continued in frame into the ras gene sequences that code for p21. When the pL promoter was derepressed, the Escherichia coli cells harboring the fusion plasmid synthesized 23000-dalton protein, which represented more than 10 percent of the total cellular protein. This protein was chimeric and contained 14 residues, which were specified by the vector; these residues were followed by all of the amino acids that make up Ha-MuSV p21ras except for four residues at the amino-terminal end. The protein appears similar to Ha-MuSV p21ras in that it undergoes immunoprecipitation by monoclonal antibodies directed toward that protein, binds guanosine diphosphate, and is capable of autophosphorylation.