Activation and inhibition of G proteins by lipoamines.

We have previously shown that alkyl-substituted amino acid derivatives directly activate G(i/o) proteins. N-Dodecyl-N(alpha),N(epsilon)-(bis-l-lysinyl)-l-lysine amide (FUB132) is a new representative of this class of compounds with increased efficacy. Here, we characterized the molecular mechanism of action of this class of compounds. FUB132 and its predecessor FUB86 were selective receptomimetics for G(i/o) because they stimulated the guanine nucleotide exchange reaction of purified G(i/o) as documented by an increased rate of GDP release, GTP gamma S binding, and GTP hydrolysis. In contrast to the receptomimetic peptide mastoparan, stimulation of G proteins by lipoamines required the presence of neither G beta gamma-dimers nor lipids. On the contrary, G beta gamma-dimers suppressed the stimulatory effect of FUB132. The stimulation of G(i/o) by lipoamines and by mastoparan was not additive. A peptide derived from the C terminus of G alpha(o3), but not a corresponding G alpha(q)-derived peptide, quenched the FUB132-induced activation of G alpha(o). In membranes prepared from human embryonic kidney 293 cells that stably expressed the G(i/o)-coupled human A(1)-adenosine receptor, lipoamines impeded high-affinity agonist binding. In contrast, antagonist binding was not affected. We conclude that alkyl-substituted amines target a site, most likely at the C terminus of G alpha(i/o)-subunits, that is also contacted by receptors. However, because G beta gamma-dimers blunt rather than enhance their efficacy, their mechanism of action differs fundamentally from that of a receptor. Thus, despite their receptomimetic effect in vitro, alkyl-substituted amines and related polyamines are poor direct G protein activators in vivo. In the presence of G beta gamma, they rather antagonize G protein-coupled receptor signaling.