Structure of an open K _ATP channel reveals tandem PIP ₂ binding sites mediating the Kir6.2 and SUR1 regulatory...

ATP-sensitive potassium (K _ATP ) channels, composed of four pore-lining Kir6.2 subunits and four regulatory sulfonylurea receptor 1 (SUR1) subunits, control insulin secretion in pancreatic β-cells. K _ATP channel opening is stimulated by PIP ₂ and inhibited by ATP. Mutations that increase channel opening by PIP ₂ reduce ATP inhibition and cause neonatal diabetes. Although considerable evidence has indicated PIP ₂ in K _ATP channel function, previously solved open-channel structures have lacked bound PIP ₂ , and mechanisms by which PIP ₂ regulates K _ATP channels remain unresolved. Here, we report the cryoEM structure of a K _ATP channel harboring the neonatal diabetes mutation Kir6.2-Q52R, bound to natural C18:0/C20:4 long-chain PIP ₂ in an open conformation. The structure reveals two adjacent PIP ₂ molecules between SUR1 and Kir6.2. The first PIP ₂ binding site is conserved among PIP ₂ -gated Kir channels. The second site forms uniquely in K _ATP at the interface of Kir6.2 and SUR1. Functional studies demonstrate both binding sites determine channel activity. Kir6.2 pore opening is associated with a twist of the Kir6.2 cytoplasmic domain and a rotation of the N-terminal transmembrane domain of SUR1, which widens the inhibitory ATP binding pocket to disfavor ATP binding. The open conformation is particularly stabilized by the Kir6.2-Q52R residue through cation-π bonding with SUR1-W51. Together, these results uncover the cooperation between SUR1 and Kir6.2 in PIP ₂ binding and gating, explain the antagonistic regulation of K _ATP channels by PIP ₂ and ATP, and provide the mechanism by which Kir6.2-Q52R stabilizes an open channel to cause neonatal diabetes.