Chimeric Galpha(q) mutants harboring the last five carboxy-terminal residues of Galpha(i2) or Galpha(o) are resistant to pertussis toxin-catalyzed ADP-ribosylation.
Three widely-used Galpha(q) chimeras harboring the last five residues of Galpha(i), Galpha(o) and Galpha(z) (qi5, qo5 and qz5) were examined for their ability to serve as substrates for pertussis toxin (PTX)-catalyzed ADP-ribosylation. In COS-7 cells coexpressing one of the three opioid receptors (mu, delta, and kappa) and a Galpha(q) chimera, agonist-induced stimulation of phosphoinositide-specific phospholipase C (PI-PLC) was largely insensitive to PTX treatment. Only the qi5-mediated stimulation of PI-PLC by kappa-opioids was partially inhibited by PTX. In betagamma-release assays, PTX treatment did not affect the ability of opioid receptors to activate these chimeras. [32P]ADP-ribosylation labeled Galpha(i/o) but not qi5 or qo5, although the expression of these chimeras was confirmed by immunodetection. Thus, Galpha(q) chimeras with a Galpha(i/o)-like tail are insensitive to PTX treatment.
Joshi
S A
SA
Department of Biology and the Biotechnology Research Institute, Hong Kong University of Science and Technology, Kowloon, China.
Fan
K P
KP
Ho
V W
VW
Wong
Y H
YH
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D000818:Animals; D019556:COS Cells; D000242:Cyclic AMP / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D007295:Inositol Phosphates / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D009249:NADP / Q000378:metabolism; D037342:Pertussis Toxin; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D051381:Rats; D017465:Receptors, Opioid, delta / Q000235:genetics / Q000502:physiology*; D017473:Receptors, Opioid, kappa / Q000235:genetics / Q000502:physiology*; D017450:Receptors, Opioid, mu / Q000235:genetics / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D007295:Inositol Phosphates; D008565:Membrane Proteins; D017465:Receptors, Opioid, delta; D017473:Receptors, Opioid, kappa; D017450:Receptors, Opioid, mu; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D010566:Virulence Factors, Bordetella; D000246:Adenosine Diphosphate Ribose; D009249:NADP; D000242:Cyclic AMP; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D054801:Phosphoinositide Phospholipase C; D019204:GTP-Binding Proteins; D043265:Phosphatidylinositol Diacylglycerol-Lyase