Kir6.2‐dependent high‐affinity repaglinide binding to β ‐cell K ATP channels

• Published in: British journal of pharmacology 2005-02, Vol.144 (4), p.551-557
• Main Authors: Hansen, Ann Maria K, Hansen, John Bondo, Carr, Richard D, Ashcroft, Frances M, Wahl, Philip
• Format: Article
• Language: English
• ISSN: 0007-1188; 1476-5381
• DOI: 10.1038/sj.bjp.0706082

The β ‐cell K ATP channel is composed of two types of subunit – the inward rectifier K + channel (Kir6.2) which forms the channel pore, and the sulphonylurea receptor (SUR1), which serves as a regulatory subunit. The N‐terminus of Kir6.2 is involved in transduction of sulphonylurea binding into channel closure, and deletion of the N‐terminus (Kir6.2ΔN14) results in functional uncoupling of the two subunits. In this study, we investigate the interaction of the hypoglycaemic agents repaglinide and glibenclamide with SUR1 and the effect of Kir6.2 on this interaction. We further explore how the binding properties of repaglinide and glibenclamide are affected by functional uncoupling of SUR1 and Kir6.2 in Kir6.2ΔN14/SUR1 channels. All binding experiments are performed on membranes in ATP‐free buffer at 37°C. Repaglinide was found to bind with low affinity ( K D =59±16 n M ) to SUR1 alone, but with high affinity (increased ∼150‐fold) when SUR1 was co‐expressed with Kir6.2 ( K D =0.42±0.03 n M ). Glibenclamide, tolbutamide and nateglinide all bound with marginally lower affinity to SUR1 than to Kir6.2/SUR1. Repaglinide bound with low affinity ( K D =51±23 n M ) to SUR1 co‐expressed with Kir6.2ΔN14. In contrast, the affinity for glibenclamide, tolbutamide and nateglinide was only mildly changed as compared to wild‐type channels. In whole‐cell patch‐clamp experiments inhibition of Kir6.2ΔN14/SUR1 currents by both repaglinide and nateglinde is abolished. The results suggest that Kir6.2 causes a conformational change in SUR1 required for high‐affinity repaglinide binding, or that the high‐affinity repaglinide‐binding site includes contributions from both SUR1 and Kir6.2. Glibenclamide, tolbutamide and nateglinide binding appear to involve only SUR1. British Journal of Pharmacology (2005) 144 , 551–557. doi: 10.1038/sj.bjp.0706082