Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.| Volume 259, ISSUE 22, P13806-13813, November 25, 1984

Isolation of two proteins with high affinity for guanine nucleotides from membranes of bovine brain.

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      Membranes from bovine brain bind relatively large quantities of guanosine 5'-(3-O-thio)triphosphate (GTP gamma S) with high affinity. The two proteins responsible for most of this activity were purified; they account for 1.5% of the membrane protein. The two proteins contain alpha subunits of either 39,000 or 41,000 Da, beta subunits of 36,000 or 35,000 Da, and a potential gamma subunit (11,000 Da). These structures are the same as a family of proteins that includes transducin and the regulatory proteins, GS and GI, of adenylate cyclase. The 41,000- and 39,000-Da polypeptides can be ADP-ribosylated with islet-activating protein from Bordetella pertussis, bind guanine nucleotides specifically, and migrate through polyacrylamide gels with rates similar to the alpha subunits of GI and transducin, respectively. The 36,000- and 35,000-Da polypeptides are similar to the beta subunits of GI and GS. The gamma subunit is found whenever beta subunits are present. The 41,000- and 39,000-Da polypeptides (with beta and gamma) are designated, respectively, GI and GO from brain. The alpha subunit of GO was isolated without the use of ligands known to dissociate other G proteins. GO alpha binds GTP gamma S reversibly in the absence of Mg2+ and is relatively stable in cholate. This isolated alpha subunit should be of great utility in elucidating the mechanism of action of this family of GTP-binding proteins.

      REFERENCES

        • Ross E.M.
        • Gilman A.G.
        Annu. Rev. Biochem. 1980; 49: 533-564
        • Stryer L.
        • Hurley J.B.
        • Fung B.K.K.
        Curr. Top. Membr. Transp. 1981; 15: 93-108
        • Gilman A.G.
        J. Clin. Invest. 1984; 73: 1-4
        • Godchaux III, W.
        • Zimmerman W.F.
        J. Biol. Chem. 1979; 254: 7874-7884
        • Baehr W.
        • Devlin M.J.
        • Applebury M.L.
        J. Biol. Chem. 1979; 254: 11669-11677
        • Kuhn H.
        Nature (Lond.). 1980; 283: 587-589
        • Hurley J.B.
        Biochem. Biophys. Res. Commun. 1980; 92: 505-510
        • Fung B.K.K.
        • Hurley J.B.
        • Stryer L.
        Proc. Natl. Acad. Sci. U. S. A. 1981; 78: 152-156
        • Northup J.K.
        • Sternweis P.C.
        • Smigel M.D.
        • Schleifer L.S.
        • Ross E.M.
        • Gilman A.G.
        Proc. Natl. Acad. Sci. U. S. A. 1980; 77: 6516-6520
        • Sternweis P.C.
        • Northup J.K.
        • Smigel M.D.
        • Gilman A.G.
        J. Biol. Chem. 1981; 256: 11517-11526
        • Hanski E.
        • Sternweis P.C.
        • Northup J.K.
        • Dromerick A.W.
        • Gilman A.G.
        J. Biol. Chem. 1981; 256: 12911-12919
        • Hanski E.
        • Gilman A.G.
        J. Cyclic Nucleotide Res. 1982; 8: 323-336
        • Bokoch G.M.
        • Katada T.
        • Northup J.K.
        • Hewlett E.L.
        • Gilman A.G.
        J. Biol. Chem. 1983; 258: 2072-2075
        • Bokoch G.M.
        • Katada T.
        • Northup J.K.
        • Ui M.
        • Gilman A.G.
        J. Biol. Chem. 1984; 259: 3560-3567
        • Codina J.
        • Hildebrandt J.
        • Iyengar R.
        • Birnbaumer L.
        • Sekura R.D.
        • Manclark C.R.
        Proc. Natl. Acad. Sci. U. S. A. 1983; 80: 4276-4280
        • Hildebrandt J.D.
        • Codina J.
        • Risinger R.
        • Birnbaumer L.
        J. Biol. Chem. 1984; 259: 2039-2042
        • Manning D.R.
        • Gilman A.G.
        J. Biol. Chem. 1983; 258: 7059-7063
        • Gill D.M.
        • Meren R.
        Proc. Natl. Acad. Sci. U. S. A. 1978; 75: 3050-3054
        • Cassel D.
        • Pfeuffer T.
        Proc. Nat. Acad. Sci. U. S. A. 1978; 75: 2669-2673
        • Katada T.
        • Ui M.
        J. Biol. Chem. 1982; 257: 7210-7216
        • Manning D.R.
        • Fraser B.A.
        • Kahn R.A.
        • Gilman A.G.
        J. Biol. Chem. 1984; 259: 749-756
        • Abood M.E.
        • Hurley J.B.
        • Pappone M.-C.
        • Bourne H.R.
        • Stryer L.
        J. Biol. Chem. 1982; 257: 10540-10543
        • Pfeuffer T.
        J. Biol. Chem. 1977; 252: 7224-7234
        • Northup J.K.
        • Smigel M.D.
        • Sternweis P.C.
        • Gilman A.G.
        J. Biol. Chem. 1983; 258: 11369-11376
        • Northup J.K.
        • Smigel M.D.
        • Gilman A.G.
        J. Biol. Chem. 1982; 257: 11416-11423
        • Katada T.
        • Bokoch G.M.
        • Smigel M.D.
        • Ui M.
        • Gilman A.G.
        J. Biol. Chem. 1984; 259: 3586-3595
        • Northup J.K.
        • Sternweis P.C.
        • Gilman A.G.
        J. Biol. Chem. 1983; 258: 11361-11368
        • Katada T.
        • Bokoch G.M.
        • Northup J.K.
        • Ui M.
        • Gilman A.G.
        J. Biol. Chem. 1984; 259: 3568-3577
        • Ross E.M.
        • Maguire M.E.
        • Sturgill T.W.
        • Biltonen R.L.
        • Gilman A.G.
        J. Biol. Chem. 1977; 252: 5761-5775
        • Schaffner W.
        • Weissman C.
        Anal. Biochem. 1973; 56: 502-504
        • Laemmli U.K.
        Nature (Lond.). 1970; 227: 680-685
        • Wray W.
        • Boulikas T.
        • Wray V.P.
        • Hancock R.
        Anal. Biochem. 1981; 118: 197-203
        • Johnson R.A.
        • Walseth T.F.
        Adv. Cyclic Nucleotide Res. 1979; 10: 135-168
        • Shaltiel S.
        Methods Enzymol. 1974; 34: 126-140
        • Schleifer L.S.
        • Garrison J.C.
        • Sternweis P.C.
        • Northup J.K.
        • Gilman A.G.
        J. Biol. Chem. 1980; 255: 2641-2644
        • Schleifer L.S.
        • Kahn R.A.
        • Hanski E.
        • Northup J.K.
        • Sternweis P.C.
        • Gilman A.G.
        J. Biol. Chem. 1982; 257: 20-23
        • Kahn R.A.
        • Gilman A.G.
        J. Biol. Chem. 1984; 259: 6228-6234
        • Smigel M.D.
        • Northup J.K.
        • Gilman A.G.
        Recent Prog. Horm. Res. 1982; 38: 601-624