Concerted gating mechanism underlying KATP channel inhibition by ATP

K-ATP channels assemble from four regulatory SUR1 and four pore-forming K(ir)6.2 subunits. At the single-channel current level, ATP-dependent gating transitions between the active burst and the inactive interburst conformations underlie inhibition of the K-ATP channel by intracellular ATP. Previously, we identified a slow gating mutation, T171A in the K(ir)6.2 subunit, which dramatically reduces rates of burst to interburst transitions in K(ir)6.2DeltaC26 channels without SUR1 in the absence of ATP. Here, we constructed all possible mutations at position 171 in K(ir)6.2DeltaC26 channels without SUR1. Only four substitutions, 171A, 171F, 171H, and 171S, gave rise to functional channels, each increasing K-i,K-ATP for ATP inhibition by >55-fold and slowing gating to the interburst by >35-fold. Moreover, we investigated the role of individual K(ir)6.2 subunits in the gating by comparing burst to interburst transition rates of channels constructed from different combinations of slow 171A and fast T171 "wild-type" subunits. The relationship between gating transition rate and number of slow subunits is exponential, which excludes independent gating models where any one subunit is sufficient for inhibition gating. Rather, our results support mechanisms where four ATP sites independently can control a single gate formed by the concerted action of all four K(ir)6.2 subunit inner helices of the K-ATP channel.