Disruption of receptor-mediated activation of G protein by mutating a conserved arginine residue in the switch II region of the α subunit

A naturally occurring point mutation (R231H) within one of the major bg-binding surface (switch II region) on the a subunit of G(S) (a(S)) has previously been found to disrupt receptor-mediated activation of G(S). The disruption caused by mutating this conserved residue may be a general phenomenon for all a subunits. Homologous mutants of the a subunit of G(Z) [a(Z); a negative regulator of adenylyl cyclase (AC)] and G(16) (alpha(16); a stimulator of phospholipase C) were constructed and examined for receptor-mediated regulation of their corresponding effecters. The mutant a(Z)R209H cannot be fully activated by the delta-opioid receptor, as indicated by the impairment of the inhibition of a(S)-stimulated AC and beta gamma-mediated stimulation of AC type II (AC2). Similarly, the mutant a(16)R216H lost the ability to mediate receptor-induced activation of phospholipase C and AC2. The receptor coupling efficacy and promiscuity of a(16)R216H were eradicated. The mutation of the conserved arginine has no observable effect on the constitutive activities of the GTPase-deficient derivatives of both a(Z) and a(16). The a subunit of G(t1) (transducin; a(t1)) attenuated bg-mediated stimulation of AC2 by sequestrating free bg subunits, but the mutant a(t1)R204H showed reduced ability to scavenge bg-mediated AC2 activation. Presumably, mutation of the conserved arginine disrupted the subunit interactions in addition to the impairment of receptor interaction.