Comparative biochemical properties of p21 ras molecules coded for by viral and cellular ras genes

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Abstract
In earlier studies, a normal cellular gene, c-rasH-1, homologous to the v-ras oncogene of Harvey murine sarcoma virus (v-rasH) was molecularly cloned. By ligating a type c retroviral promotor to c-rasH-1, [mouse] NIH-3Te cells could be transformed with the c-rasH-1 gene. The transformed cells contained high levels of a p21 protein coded for by the c-rasH-1 gene. In the current studies, v-rasH p21 and c-rasH p21 were extensively purified and the in vivo and in vitro biochemical properties of both molecules were compared. The p21 proteins coded for by v-rasH and c-rasH-1 shared certain properties: each was synthesized as a precursor protein which subsequently became bound to the inner surface of the plasma membrane; each was associated with guanine nucleotide-binding activity, a property which copurified with p21 molecules on a high-pressure liquid chromatography molecular sizing column. In vivo, .apprx. 20-30% of v-rasH p21 molecules were in the form of phosphothreonine-containing pp21 molecules; in vivo, only a minute fraction of c-rasH-1 p21 contained phosphate which was found on a serine residue. The v-rasH pp21 molecules with an authentic phosphothreonine peptide could be synthesized in vitro in an autophosphorylation reaction in which the .gamma.-phosphate of GTP was transferred to vrasH p21. No autophosphorylating activity was associated with purified c-rasH-1 p21 in vitro. The results indicate a major qualitative difference between the p21 proteins coded for by vrasH and c-rasH-1. The p21 coded for by a mouse-derived oncogenic virus, BALB murine sarcoma virus, resembled the p21 coded for by c-rasH-1 in that it bound guanine nucleotides but did not label appreciably with 32Pi. Comparison of the forms of p21 coded for by other members of the ras gene family indicated that the guanine nucleotide-binding activity is common to p21 molecules coded for by all known members of the family.