Reinterpreting the Action of ATP Analogs on KATP Channels*
Background: Stimulation by post-hydrolytic, ADP-bound conformations of SUR1 underlies current models of KATP channel activation; ATP analogs are assumed to lower activity by reducing hydrolysis. Results: ATPγS switches conformations with lowered affinity; AMP-PxP selectively bind NBD1, preventing switching. Conclusion: The actions of ATP analogs on KATP channels require reinterpretation. Significance: Reduced affinities of SUR1 NBDs for ATP analogs limit conformational switching and channel activity. Neuroendocrine-type KATP channels, (SUR1/Kir6.2)4, couple the transmembrane flux of K+, and thus membrane potential, with cellular metabolism in various cell types including insulin-secreting β-cells. Mutant channels with reduced activity are a cause of congenital hyperinsulinism, whereas hyperactive channels are a cause of neonatal diabetes. A current regulatory model proposes that ATP hydrolysis is required to switch SUR1 into post-hydrolytic conformations able to antagonize the inhibitory action of nucleotide binding at the Kir6.2 pore, thus coupling enzymatic and channel activities. Alterations in SUR1 ATPase activity are proposed to contribute to neonatal diabetes and type 2 diabetes risk. The regulatory model is partly based on the reduced ability of ATP analogs such as adenosine 5′-(β,γ-imino)triphosphate (AMP-PNP) and adenosine 5′-O-(thiotriphosphate) (ATPγS) to stimulate channel activity, presumably by reducing hydrolysis. This study uses a substitution at the catalytic glutamate, SUR1E1507Q, with a significantly increased affinity for ATP, to probe the action of these ATP analogs on conformational switching. ATPγS, a slowly hydrolyzable analog, switches SUR1 conformations, albeit with reduced affinity. Nonhydrolyzable AMP-PNP and adenosine 5′-(β,γ-methylenetriphosphate) (AMP-PCP) alone fail to switch SUR1, but do reverse ATP-induced switching. AMP-PCP displaces 8-azido-[32P]ATP from the noncanonical NBD1 of SUR1. This is consistent with structural data on an asymmetric bacterial ABC protein that shows that AMP-PNP binds selectively to the noncanonical NBD to prevent conformational switching. The results imply that MgAMP-PNP and MgAMP-PCP (AMP-PxP) fail to activate KATP channels because they do not support NBD dimerization and conformational switching, rather than by limiting enzymatic activity.