Nucleotide inhibition of the pancreatic ATP-sensitive K+ channel explored with patch-clamp fluorometry
Abstract
Pancreatic ATP-sensitive K+ channels (KATP) comprise four inward rectifier subunits (Kir6.2), each associated with a sulphonylurea receptor (SUR1). ATP/ADP binding to Kir6.2 shuts KATP. Mg-nucleotide binding to SUR1 stimulates KATP. In the absence of Mg2+, SUR1 increases the apparent affinity for nucleotide inhibition at Kir6.2 by an unknown mechanism. We simultaneously measured channel currents and nucleotide binding to Kir6.2. Fits to combined data sets suggest that KATP closes with only one nucleotide molecule bound. A Kir6.2 mutation (C166S) that increases channel activity did not affect nucleotide binding, but greatly perturbed the ability of bound nucleotide to inhibit KATP. Mutations at position K205 in SUR1 affected both nucleotide affinity and the ability of bound nucleotide to inhibit KATP. This suggests a dual role for SUR1 in KATP inhibition, both in directly contributing to nucleotide binding and in stabilising the nucleotide-bound closed state.
Data availability
All data and associated code are available on github (https://github.com/smusher/KATP_paper_2019) and have also been uploaded to Dryad (https://doi.org/10.5061/dryad.0vt4b8gtv).
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Nucleotide inhibition of the pancreatic ATP-sensitive K+ channel explored with patch-clamp fluorometryDryad Digital Repository, doi:10.5061/dryad.0vt4b8gtv.
Article and author information
Author details
Funding
Biotechnology and Biological Sciences Research Council (BB/R002517/1)
- Frances M Ashcroft
- Michael C Puljung
John Fell Fund, University of Oxford
- Michael C Puljung
Wellcome (203731/Z/16/A)
- Samuel G Usher
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Gary Yellen, Harvard Medical School, United States
Version history
- Received: October 16, 2019
- Accepted: January 7, 2020
- Accepted Manuscript published: January 7, 2020 (version 1)
- Version of Record published: February 6, 2020 (version 2)
- Version of Record updated: February 27, 2020 (version 3)
Copyright
© 2020, Usher et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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