The proline-rich N-terminal domain of G18 exhibits a novel G protein regulatory function.
The protein G18 (also known as AGS4 or GPSM3) contains three conserved GoLoco/GPR domains in its central and C-terminal regions that bind to inactive Galpha(i), whereas the N-terminal region has not been previously characterized. We investigated whether this domain might itself regulate G protein activity by assessing the abilities of G18 and mutants thereof to modulate the nucleotide binding and hydrolytic properties of Galpha(i1) and Galpha(o). Surprisingly, in the presence of fluoroaluminate (AlF(4)(-)) both G proteins bound strongly to full-length G18 (G18wt) and to its isolated N-terminal domain (G18DeltaC) but not to its GoLoco region (DeltaNG18). Thus, it appears that its N-terminal domain promotes G18 binding to fluoroaluminate-activated Galpha(i/o). Neither G18wt nor any G18 mutant affected the GTPase activity of Galpha(i1) or Galpha(o). In contrast, complex effects were noted with respect to nucleotide binding. As inferred by the binding of [(35)S]GTPgammaS (guanosine 5'-O-[gamma-thio]triphosphate) to Galpha(i1), the isolated GoLoco region as expected acted as a guanine nucleotide dissociation inhibitor, whereas the N-terminal region exhibited a previously unknown guanine nucleotide exchange factor effect on this G protein. On the other hand, the N terminus inhibited [(35)S]GTPgammaS binding to Galpha(o), albeit to a lesser extent than the effect of the GoLoco region on Galpha(i1). Taken together, our results identify the N-terminal region of G18 as a novel G protein-interacting domain that may have distinct regulatory effects within the G(i/o) subfamily, and thus, it could potentially play a role in differentiating signals between these related G proteins.
Zhao
Peishen
P
Department of Physiology and Pharmacology, The University of Western Ontario, London, Ontario N6A 5C1, Canada.
Nguyen
Chau H
CH
Chidiac
Peter
P
D000595:Amino Acid Sequence; D000818:Animals; D000939:Epitopes / Q000737:chemistry; D019204:GTP-Binding Proteins / Q000737:chemistry*; D020730:Guanine Nucleotide Dissociation Inhibitors / Q000737:chemistry* / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000737:chemistry; D006160:Guanosine Triphosphate / Q000737:chemistry; D006868:Hydrolysis; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D011392:Proline / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D013997:Time Factors; D014018:Tissue Distribution
D000939:Epitopes; C548537:G18 protein, mouse; D020730:Guanine Nucleotide Dissociation Inhibitors; D011994:Recombinant Proteins; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D011392:Proline; D019204:GTP-Binding Proteins