MgADP Antagonism to Mg2+-independent ATP Binding of the Sulfonylurea Receptor SUR1

Kazumitsu Ueda,Nobuya Inagaki,Susumu Seino

doi:10.1074/jbc.272.37.22983

Abstract

Pancreatic beta-cell ATP-sensitive potassium (KATP) channels play an important role in the regulation of glucose-induced insulin secretion. The beta-cell KATP channel comprises two subunits, the sulfonylurea receptor SUR1, a member of the ATP-binding cassette (ABC) superfamily, and Kir6.2, a member of the inward rectifier K+ channel family. The activity of the KATP channel is under complex regulation by the intracellular ATP and ADP. To understand the roles of the two nucleotide-binding folds (NBFs) of SUR1 in the regulation of KATP channel activity, we introduced point mutations into the core consensus sequence of the Walker A or B motif of each NBF of SUR1 and characterized ATP binding and ADP or MgADP antagonism to it. SUR1 was efficiently photolabeled with 8-azido-[alpha-32P]ATP and 8-azido-[gamma-32P]ATP in the presence or absence of Mg2+ or vanadate. NBF1 mutations impaired ATP binding, but NBF2 mutations did not. MgADP strongly antagonized ATP binding, and the NBF2 mutation reduced MgADP antagonism. These results show that SUR1, unlike other ABC proteins, strongly binds ATP at NBF1 even in the absence of Mg2+ and that MgADP, through binding at NBF2, antagonizes the Mg2+-independent high affinity ATP binding at NBF1.

Highlights

Pancreatic ␤-cell ATP-sensitive potassium (KATP) channels play an important role in the regulation of glucose-induced insulin secretion
We show in the present study that unlike other ATPbinding cassette (ABC) proteins such as Pgp and multidrug resistance-associated protein (MRP), SUR1 strongly and stably binds ATP at NBF1 even in the absence of Mg2ϩ, and that MgADP, through binding at NBF2, antagonizes the Mg2ϩ-independent high affinity ATP binding at NBF1
SUR1—We investigated whether SUR1 trapped a nucleotide at the catalytic site in the presence of vanadate, as has been reported of Pgp

Summary

Introduction

Pancreatic ␤-cell ATP-sensitive potassium (KATP) channels play an important role in the regulation of glucose-induced insulin secretion. We show in the present study that unlike other ABC proteins such as Pgp and MRP, SUR1 strongly and stably binds ATP at NBF1 even in the absence of Mg2ϩ, and that MgADP, through binding at NBF2, antagonizes the Mg2ϩ-independent high affinity ATP binding at NBF1.

Results

Conclusion