An intact n terminus of the gamma subunit is required for the G beta gamma stimulation of rhodopsin phosphorylation by human beta-adrenergic receptor kinase-1 but not for kinase binding

Cleavage after lysine 32 in the G gamma(2) subtype and after lysine 36 in the G gamma(3) subtype of purified mixed brain G beta gamma by endoproteinase Lys-C blocks G beta gamma-mediated stimulation of phosphorylation of rhodopsin in urea-extracted rod outer segments by recombinant human beta-adrenergic receptor kinase (h beta ARK1) holoenzyme while h beta ARK1 binding to rod outer segments is partially affected. This treatment does not attenuate the binding of the treated G beta gamma to C-terminal fragments of h beta ARK1 containing the pleckstrin homology domain. Lys-C proteolysis also does not alter the association of the G beta gamma with phospholipids, its ability to support pertussis toxin-catalyzed G alpha(0)/G alpha(i) ADP-ribosylation, or its ability to inhibit forskolin stimulated platelet adenylate cyclase. The G beta subunit remains noncovalently associated with the cleaved G gamma fragments. Thus, in addition to recruiting h beta ARK1 to its receptor substrate, G gamma contributes secondary and/or tertiary structural features to activate the kinase.