Hydrogen-bonding dynamics between adjacent blades in G-protein beta-subunit regulates GIRK channel activation.
Functionally critical domains in the betagamma-subunits of the G-protein (Gbetagamma) do not undergo large structural rearrangements upon binding to other proteins. Here we show that a region containing Ser(67) and Asp(323) of Gbetagamma is a critical determinant of G-protein-gated inwardly rectifying K(+) (GIRK) channel activation and undergoes only small structural changes upon mutation of these residues. Using an interactive experimental and computational approach, we show that mutants that form a hydrogen-bond between positions 67 and 323 do not activate a GIRK channel. We also show that in the absence of hydrogen-bonding between these positions, other factors, such as the displacement of the crucial Ggamma residues Pro(60) and Phe(61), can impair Gbetagamma-mediated GIRK channel activation. Our results imply that the dynamic nature of the hydrogen-bonding pattern in the wild-type serves an important functional role that regulates GIRK channel activation by Gbetagamma and that subtle changes in the flexibility of critical domains could have substantial functional consequences. Our results further strengthen the notion that the dynamic regulation of multiple interactions between Gbetagamma and effectors provides for a complex regulatory process in cellular functions.
Mirshahi
Tooraj
T
Department of Physiology and Biophysics, Mount Sinai School of Medicine, New York, New York 10029, USA.
Logothetis
Diomedes E
DE
Rosenhouse-Dantsker
Avia
A
D000818:Animals; D003198:Computer Simulation; D019281:Dimerization; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D044387:GTP-Binding Protein beta Subunits / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D044388:GTP-Binding Protein gamma Subunits / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D006860:Hydrogen Bonding; D066298:In Vitro Techniques; D015640:Ion Channel Gating; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D011485:Protein Binding; D011487:Protein Conformation; D014982:Xenopus laevis
D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits