Location of the  4 Transmembrane Helices in the BK Potassium Channel (original) (raw)

2009, Journal of Neuroscience

Large-conductance, voltage-and Ca 2ϩ-gated potassium (BK) channels control excitability in a number of cell types. BK channels are composed of ␣ subunits, which contain the voltage-sensor domains and the Ca 2ϩ-sensor domains and form the pore, and often one of four types of ␤ subunits, which modulate the channel in a cell-specific manner. ␤4 is expressed in neurons throughout the brain. Deletion of ␤4 in mice causes temporal lobe epilepsy. Compared with channels composed of ␣ alone, channels composed of ␣ and ␤4 activate and deactivate more slowly. We inferred the locations of the two ␤4 transmembrane (TM) helices TM1 and TM2 relative to the seven ␣ TM helices, S0-S6, from the extent of disulfide bond formation between cysteines substituted in the extracellular flanks of these TM helices. We found that ␤4 TM2 is close to ␣ S0 and that ␤4 TM1 is close to both ␣ S1 and S2. At least at their extracellular ends, TM1 and TM2 are not close to S3-S6. In six of eight of the most highly crosslinked cysteine pairs, four crosslinks from TM2 to S0 and one each from TM1 to S1 and S2 had small effects on the V 50 and on the rates of activation and deactivation. That disulfide crosslinking caused only small functional perturbations is consistent with the proximity of the extracellular ends of TM2 to S0 and of TM1 to S1 and to S2, in both the open and closed states. Materials and Methods Constructs. Mutants of the BK ␣ subunit (mSlo1, KCNMA1; GenBank accession number NM_010610; 1169 residues; molecular weight 131,700) and human BK ␤4 subunit (KCNMB4, Open Biosystems clone/