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Hydrogen-Bonding Dynamics between Adjacent Blades in G-Protein ß-Subunit Regulates GIRK Channel Activation

Department of Physiology and Biophysics, Mount Sinai School of Medicine, New York, New York 10029

Correspondence: Address reprint requests to Avia Rosenhouse-Dantsker, E-mail: avia.rosenhouse-dantsker{at}mssm.edu.

Functionally critical domains in the ß-subunits of the G-protein (Gß) do not undergo large structural rearrangements upon binding to other proteins. Here we show that a region containing Ser⁶⁷ and Asp³²³ of Gß 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 G residues Pro⁶⁰ and Phe⁶¹, can impair Gß-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 Gß 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 Gß and effectors provides for a complex regulatory process in cellular functions.