Binding of regulator of G protein signaling (RGS) proteins to phospholipidbilayers - Contribution of location and/or orientation to GTPase-activating protein activty

Regulator of G protein signaling (RGS) proteins must bind membranes in an o rientation that permits the protein-protein interactions necessary for regu latory activity. RGS4 binds to phospholipid surfaces in a slow, multistep p rocess that leads to maximal GTPase-activating protein (GAP) activity. When RGS4 is added to phospholipid vesicles that contain m2 or m1 muscarinic re ceptor and G(i), G(z), or G(q), GAP activity increases similar to3-fold ove r 4 h at 30 degreesC and more slowly at 20 degreesC, This increase in GAP a ctivity is preceded by several other events that suggest that, after bindin g, optimal interaction with G protein and receptor requires reorientation o f RGS4 on the membrane surface, a conformational change, or both. Binding o f RGS4 is initially reversible but becomes irreversible within 5 min. Onset of irreversibility parallels initial quenching of tryptophan fluorescence (t(1/2) similar to 30 s), Further quenching occurs after binding has become irreversible (t(1/2) similar to 6 min) but is complete well before maximal GAP activity is attained. These processes all appear to be energetically d riven by the amphipathic N-terminal domain of RGS4 and are accelerated by p almitoylation of cysteine residues in this region. The RGS4 N-terminal doma in confers similar membrane binding behavior on the RGS domains of either R GS10 or RGSZ1.