GTP analogues promote release of the α subunit of the guanine nucleotide binding protein, G(i)2, from membranes of rat glioma C6 BU1 cells

The major pertussis-toxin-sensitive guanine nucleotide-binding protein of rat glioma C6 BU1 cells corresponded immunologically to G(i)2. Antibodies which recognize the α subunit of this protein indicated that it has an apparent molecular mass of 40 kDa and a pI of 5.7. Incubation of membranes of these cells with guanosine 5'-[βγ-imido]triphosphate, or other analogues of GTP, caused release of this polypeptide from the membrane in a time-dependent manner. Analogues of GDP or of ATP did not mimic this effect. The GTP analogues similarly caused release of the α subunit of G(i)2 from membranes of C6 cells in which this G-protein had been inactivated by pretreatment with pertussis toxin. The β subunit was not released from the membrane under any of these conditions, indicating that the release process was a specific response to the dissociation of the G-protein after binding of the GTP analogue. Similar nucleotide profiles for release of the α subunits of forms of G(i) were noted for membranes of both the neuroblastoma x glioma hybrid cell line NG108-15 and of human platelets. These data provide evidence that: (a) pertussis-toxin-sensitive G-proteins, in native membranes, do indeed dissociate into α and βγ subunits upon activation; (2) the α subunit of 'G₁-like' proteins need not always remain in intimate association with the plasma membrane; and (3) the α subunit of G(i)2 can still dissociate from the β/γ subunits after pertussis-toxin-catalyzed ADP-ribosylation.