Binding of the G protein βγ subunit to multiple regions of G protein‐gated inward‐rectifying K⁺ channels

<jats:p>We have previously shown that direct binding of the βγ subunit of G protein (Gβγ) to both the N‐terminal domain and the C‐terminal domain of a cloned G protein‐gated inward‐rectifying K<jats:sup>+</jats:sup> channel subunit, GIRK1, is important for channel activation. We have now further localized the Gβγ binding region in the N‐terminal domain of GIRK1 to amino acids 34–86 and the Gβγ binding region in the C‐terminal domain of GIRK1 to two separate fragments of amino acids 318–374 and amino acids 390–462. Of the four cloned mammalian GIRK subunits, GIRK1–4, GIRK1 and 4 form heteromeric K<jats:sup>+</jats:sup> channels in the heart and similar channels in the brain include heteromultimers of GIRK1 and 2, and possibly other GIRK homomultimers and heteromultimers. We found that the N‐terminal and the C‐terminal domains of all four GIRKs bound Gβγ. The Gβγ binding activities for the C‐terminal domains of GIRK2–4 were lower than that for the C‐terminal domain of GIRK1. The higher Gβγ binding activity for the C‐terminal domain of GIRK1 is due to amino acids 390–462 which are unique to GIRK1. We also found that the N‐terminal and C‐terminal domains of GIRKs interacted with each other, and the N‐terminal domain of either GIRK1 or GIRK4 together with the C‐terminal domain of GIRK1 exhibited much enhanced binding of Gβγ. These results are consistent with the idea that the N‐ and C‐terminal domains of the cardiac G protein‐gated K<jats:sup>+</jats:sup> channel subunits may interact with each other to form higher affinity binding site(s) for Gβγ.</jats:p>