J Physiol Volume 584, Number 3, 743-753, November 1, 2007 DOI: 10.1113/jphysiol.2007.143149
A mutation in the ATP-binding site of the Kir6.2 subunit of the KATP channel alters coupling with the SUR2A subunit
Paolo Tammaro1 and
Frances M. Ashcroft1
1 Department of Physiology, Anatomy and Genetics, Oxford University, Oxford OX1 3PT, UK
Mutations in the pore-forming subunit of the ATP-sensitive K+ (KATP) channel Kir6.2 cause neonatal diabetes. Understanding the molecular mechanism of action of these mutations has provided valuable insight into the relationship between the structure and function of the KATP channel. When Kir6.2 containing a mutation (F333I) in the putative ATP-binding site is coexpressed with the cardiac type of regulatory KATP channel subunit, SUR2A, the channel sensitivity to ATP inhibition is reduced and the intrinsic open probability (Po) is increased. However, the extent of macroscopic current activation by MgADP was unaffected. Here we examine rundown and MgADP activation of wild-type and Kir6.2-F333I/SUR2A channels using single-channel recording, noise analysis and spectral analysis. We also compare the effect of mutating the adjacent residue, G334, on rundown and MgADP activation. All three approaches indicated that rundown of Kir6.2-F333I/SUR2A channels is due to a reduction in the number of active channels in the patch and that MgADP reactivation involves recruitment of inactive channels. In contrast, rundown and MgADP reactivation of wild-type and Kir6.2-G334D/SUR2A channels, and of Kir6.2-F333I/SUR1 channels, involve a gradual change in Po. Our results suggest that F333 in Kir6.2 interacts functionally with SUR2A to modulate channel rundown and MgADP activation. This interaction is fairly specific as it is not disturbed when the adjacent residue (G334) is mutated. It is also not a consequence of the enhanced Po of Kir6.2-F333I/SUR2A channels, as it is not found for other mutant channels with high Po (Kir6.2-I296L/SUR2A).
(Received 15 August 2007;
accepted after revision 6 September 2007;
first published online 13 September 2007)
Corresponding author F. M. Ashcroft: Department of Physiology, Anatomy and Genetics, Parks Road, Oxford OX1 3PT, UK. Email: frances.ashcroft{at}physiol.ox.ac.uk
This paper has online supplemental material.
Copyright © 2007 The Physiological Society.
