Structure and dynamics of the pore of inwardly rectifying K-ATP channels

Citation
G. Loussouarn et al., Structure and dynamics of the pore of inwardly rectifying K-ATP channels, J BIOL CHEM, 275(2), 2000, pp. 1137-1144
Citations number
41
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Biochemistry & Biophysics
Journal title
JOURNAL OF BIOLOGICAL CHEMISTRY
ISSN journal
0021-9258 → ACNP
Volume
275
Issue
2
Year of publication
2000
Pages
1137 - 1144
Database
ISI
SICI code
0021-9258(20000114)275:2<1137:SADOTP>2.0.ZU;2-I
Abstract
Inwardly rectifying K+ currents are generated by a complex of four Kir (Kir 1-6) subunits. Pore properties are conferred by the second transmembrane do main (M2) of each subunit. Using cadmium ions as a cysteine-interacting pro be, we examined the accessibility of substituted cysteines in M2 of the Kir 6.2 subunit of inwardly rectifying K-ATP channels. The ability of Cd2+ ions to inhibit channels was used as the estimate of accessibility. The distrib ution of Cd2+ accessibility is consistent with an alpha-helical structure o f M2. The apparent surface of reactivity is broad, and the most reactive re sidues correspond to the solvent-accessible residues in the bacterial KcsA channel crystal structure. In several mutants, single channel measurements indicated that inhibition occurred by a single transition from the open sta te to a zero-conductance state. Analysis of currents expressed from mixture s of control and L164C mutant subunits indicated that at least three cystei nes are required for coordination of the Cd2+ ion. Application of phosphati dylinositol 4,Ei-diphosphate to inside-out membrane patches stabilized the open state of all mutants and also reduced cadmium sensitivity, Moreover, t he Cd2+ sensitivity of several mutants was greatly reduced in the presence of inhibitory ATP concentrations. Taken together, these results are consist ent with state-dependent accessibility of single Cd2+ ions to coordination sites within a relatively narrow inner vestibule.