Role for SUR 2 A ED Domain in Allosteric Coupling within the K ATP Channel Complex

Adoption of different functional states in response to effector binding at a site distal from the catalytic site within homomeric systems, with one or more axes of molecular symmetry, defi nes the allosteric regulation of enzymes ( Monod et al., 1965 ; Monod, 1966 ). This is the case with ATP-binding cassette (ABC) transporters where adenine nucleotide interactions with nucleotide binding domains (NBDs) drive structural rearrangement necessary for cargo translocation across the cell membrane ( Linton and Higgins, 2007 ). Within the ABC superfamily, regulation of the ATP-sensitive K + (K ATP ) channel complex is unique as it relies not only on structural coupling and cooperativity, induced by nucleotide interaction within the ATPaseharboring regulatory sulfonylurea receptors (SUR), but also on transmission of such interaction to the associated pore-forming Kir6.x channel ( Alekseev et al., 2005 ). The K ATP channel heteromer is comprised of four SUR modules, members of the ABCC subfamily, and four K + conducting pores ( Aguilar-Bryan et al., 1995 ; Inagaki et al., 1995; 1996 ; Tucker et al., 1997 ; Lorenz and Terzic, 1999 ; Nichols, 2006 ). SUR, like other ABCC proteins, consists of two bundles of six hydrophobic transmembrane-spanning domains (TMDs) fused to hydrophilic nucleotide-binding domains (NBDs) (TMD 1 -NBD1-TMD 2 NBD2) ( Lorkowski and Cullen, 2002 ). An additional TMD 0 module with fi ve transmembrane domains (TMD 0 TMD 1 -NBD1-TMD 2 -NBD2) anchors the channel pore to the SUR module ( Babenko and Bryan, 2003 ; Fang Abbreviations used in this paper: ABC, ATP-binding cassette; CL, cytoplasmic loop; HRP, horseradish peroxidase; KCO, potassium channel opener; NBD, nucleotide binding domain; SUR, sulfonylurea receptor; TMD, transmembrane-spanning domain. et al., 2006 ; Hosy et al., 2007 ). While SUR shares the properties of ATP interaction and hydrolysis with ABC proteins ( Bienengraeber et al., 2000; Matsuo et al., 2000 ), a catalysis-coupled transport function has not been identifi ed for K ATP channels. Catalysis in SUR, while not required for ion permeation down the electrochemical gradient, has been implicated in allosteric regulation of the heteromeric K ATP channel complex ( Zingman et al., 2001 ). In this way, the concept of allosteric regulation, developed for homomers, could be expanded to heteromers, where ligand-induced conformation adaptation within the regulatory module is translated to the functional state of the distal subunit. While the allosteric hierarchy controlling K ATP channel operation has not been dissected, the proposed model of nucleotide-dependent channel gating ( Abraham et al., 2002 ; Selivanov et al., 2004 ) implies that four identical binding sites for ATP and ATP/ADP coexist on the poreforming and regulatory subunits, respectively, within the octameric complex ( Clement et al., 1997 ). Binding of ATP to the Kir6.x pore inhibits channel opening ( Tucker et al., 1997 ; Drain et al., 1998 ), whereas MgADP at the corresponding regulatory SUR subunit antagonizes ATP-induced pore closure ( Shyng et al., 1997 ; Matsuo et al., 2000 ). Cooperative rather than individual contribution of SUR NBDs gate channel operation as Role for SUR2A ED Domain in Allosteric Coupling within the K ATP Channel Complex