Purification of a protein cofactor required for ADP-ribosylation of the stimulatory regulatory component of adenylate cyclase by cholera toxin.
A factor (ARF) that is required for the cholera toxin-dependent ADP-ribosylation of the stimulatory, GTP-binding regulatory component (Gs) of adenylate cyclase has been purified about 2000-fold from cholate extracts of rabbit liver membranes. ARF is an intrinsic membrane protein with Mr = 21,000. The final product can be resolved into two polypeptides with very similar molecular weights; each of these has ARF activity. The ADP-ribosylation of Gs can now be studied with defined components. GTP and ARF are both necessary cofactors. The data imply that the substrates for the activated toxin are NAD and a GTP X Gs X ARF complex, and the reaction proceeds in a lipid environment. The apparent ability of ARF to bind to the alpha subunit of Gs suggests that it may play another, unknown role in the regulation of adenylate cyclase activity.
D020727:ADP-Ribosylation Factors; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D002462:Cell Membrane / Q000201:enzymology; D002772:Cholera Toxin / Q000494:pharmacology*; D019204:GTP-Binding Proteins; D006160:Guanosine Triphosphate / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008565:Membrane Proteins / Q000302:isolation & purification* / Q000502:physiology; D008970:Molecular Weight; D009702:Nucleoside Diphosphate Sugars / Q000378:metabolism*; D011485:Protein Binding; D011817:Rabbits; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism*
D008565:Membrane Proteins; D009702:Nucleoside Diphosphate Sugars; D011956:Receptors, Cell Surface; D000246:Adenosine Diphosphate Ribose; D006160:Guanosine Triphosphate; D002772:Cholera Toxin; D019204:GTP-Binding Proteins; D020727:ADP-Ribosylation Factors; D000262:Adenylyl Cyclases