Interaction between Pleckstrin Homology Domains and G Protein βγ-Subunits : Analyses of Kinetic Parameters by a Biosensor-Based Method

抄録

Pleckstrin homology (PH) domains, comprised of rather weakly conserved sequences of about 100 amino acid residues, are a protein motif found in many signaling and cytoskeletal proteins. PH domains have been shown to bind to the βγ subunits of heterotrimeric GTP-binding proteins (G_βγ), but the affinity of PH domains for G_βγ has not been quantitatively estimated in detail. To characterize the nature of the interaction between PH domains and G_βγ, its kinetic parameters were analyzed using a BIAcore^TM instrument. All PH domains tested (PH domains of ras-specific guanine nucleotide exchange factor (ras-GRF), phospholipase (PLC) γ1, and Son of sevenless protein (Sos)) appeared to bind to G_β1γ2 with affinity constants K_D of 0.108, 0.318, and 0.208 μM, respectively. The binding of PH domains to G_βγ was inhibited by preincubation of G_βγ with the GDP-bound but not the GTP-bound form of Gi_α. This study showed a high affinity interaction between PH domains and G_βγ and suggests a potential role of PH domains in G_βγ-mediated signal transduction in intact cells.