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Involvement of the N‐terminus of Kir6.2 in the inhibition of the K ATP channel by ATP
Author(s) -
Proks Peter,
Gribble Fiona M.,
Adhikari Raju,
Tucker Stephen J.,
Ashcroft Frances M.
Publication year - 1999
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1111/j.1469-7793.1999.019af.x
Subject(s) - kir6.2 , atp sensitive potassium channel , n terminus , potassium channel , chemistry , channel (broadcasting) , adenosine triphosphate , biophysics , microbiology and biotechnology , biochemistry , biology , endocrinology , protein subunit , glibenclamide , computer science , gene , computer network , diabetes mellitus , peptide sequence
1 ATP‐sensitive potassium (K ATP ) channels are composed of pore‐forming Kir6.2 and regulatory SUR subunits. A truncated isoform of Kir6.2, Kir6.2ΔC26, expresses ATP‐sensitive channels in the absence of SUR1, suggesting the ATP‐inhibitory site lies on the Kir6.2 subunit. 2 We examined the effect on the channel ATP sensitivity of mutating the arginine residue at position 50 (R50) in the N‐terminus of Kir6.2, by recording macroscopic currents in membrane patches excised from Xenopus oocytes expressing wild‐type or mutant Kir6.2ΔC26. 3 Substitution of R50 by serine, alanine or glycine reduced the K i for ATP inhibition from 117 μ m to 800 μ m , 1.1 m m and 3.8 m m , respectively. The single‐channel conductance and kinetics were unaffected by any of these mutations. Mutation to glutamate, lysine, asparagine, glutamine or leucine had a smaller effect ( K i , ∼300–400 μ m ). The results indicate that the side chain of the arginine residue at position 50 is unlikely to contribute directly to the binding site for ATP, and suggest it may affect ATP inhibition by allosteric interactions. 4 Mutation of the isoleucine residue at position 49 to glycine (I49G) reduced the channel ATP sensitivity, while the mutation of the glutamate residue at position 51 to glycine (E51G) did not. 5 When a mutation in the N‐terminus of Kir6.2ΔC26 that alters ATP sensitivity (R50S; K i , 800 μ m ) was combined with one in the C‐terminus (E179Q; K i , 300 μ m ), the K i for the apparent ATP sensitivity was increased to 2.8 m m . The Hill coefficient was also increased. This suggests that the N‐ and C‐termini of Kir6.2 may co‐operate to influence channel closure by ATP.