IMMUNOAFFINITY PURIFICATION AND CHARACTERIZATION OF THE ALPHA-SUBUNITS OF G0 TYPE-G PROTEINS FROM VARIOUS BOVINE-TISSUES

Polyclonal antibodies to the alpha-subunits of G(o) type G proteins (G(o)alpha) were coupled to agarose gel and used to isolate G(o)alpha from solubilized membranes of various bovine tissues. The cholate extract of membranes was applied to the anti-G(o)alpha-agarose gel column. The column was washed extensively, then bound proteins were eluted at a neutral pH using a commercial ActiSep Elution Medium. The proteins in the eluate displayed a single band of 39 kDa on SDS-polyacrylamide gel electrophoresis. They bound to GTP-gamma-S and were ADP-ribosylated by pertussis toxin. The yield of the immunoreactive G(o)alpha from the extract was about 40%. Isoelectric focusing, immunoassay and peptide mapping analysis of the G(o)alpha-like proteins purified from the heart and adrenal medulla indicated that these proteins were very similar to the alpha subunit of a minor subtype of G(o) in the brain which was previously referred to as G(o)*alpha.