Purification of unique α subunits of GTP-binding regulatory proteins (G proteins) by affinity chromatography with immobilized βγ subunits

Iok Hou Pang, Paul C. Sternweis

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121 Scopus citations

Abstract

Novel G protein α subunits were purified from rat brain by an affinity matrix containing immobilized βγ subunits (Pang, I.-H., and Sternweis, P.C. (1989) Proc. Natl. Acad. Sci. U.S.A. 86, 7814-7818). They were unique based on the following criteria. These α subunits migrated differently through polyacrylamide gels with an apparent molecular mass of 42 kDa. They did not behave similarly to the other brain G proteins by conventional chromatographic techniques. Antisera raised against a common region of known α subunits failed to recognize these 42-kDa polypeptides. Finally, primary sequences of tryptic fragments of these proteins contain regions homologous to, yet unique from, the other α subunits. Sequences are identical with one or more members of a new family of α subunits recently identified by molecular genetic techniques (Strathmann, M., Wilke, T.M., and Simon, M.I. (1989) Proc. Natl. Acad. Sci. U.S.A. 86, 7407-7409); most of the primary sequence identifies an α subunit labeled α(q). These polypeptides were not substrates for ADP-ribosylation catalyzed by pertussis toxin. They bound GTPγS only with slow rates and low stoichiometry. Antisera to peptides based on primary sequence were specific for the new α subunits and indicate that they are widely distributed at low levels in different tissues but more concentrated in brain and lung. This procedure provides a means of preparing native G proteins that have a potential role as modulators of pertussis toxin-insensitive regulatory pathways.

Original languageEnglish (US)
Pages (from-to)18707-18712
Number of pages6
JournalJournal of Biological Chemistry
Volume265
Issue number30
StatePublished - Nov 30 1990

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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