Beta Subunits Bridge Two Alpha Subunits Within the BK Channel Tetramer

Abstract

BK potassium channels contain four pore-forming alpha subunits and four modulatory beta subunits. Transmembrane (TM) helices S1-S6 of BK alpha are homologous to S1-S6 of other V-gated potassium channels; however, BK alpha contains a unique seventh TM helix, S0, N terminal to S1. The beta subunits contain two TM helices, TM1 and TM2. From the extent of endogenous disulfide crosslinking between Cys substituted for the four residues just flanking the extracellular ends of alpha S0-S6 and of beta1 TM1 and TM2, we previously inferred that the flank of S0 was closest to the four-residue loop between S3 and S4 and also contacted the flanks of S1 and S2. Furthermore, the flank of beta1 TM1 was closest to the flanks of S1 and S2, and the flank of TM2 was closest to the flank of S0. We have now extended this analysis to the membrane doman. We find that Cys in the first helical turn of S0 within the membrane forms disulfides with Cys subsituted in the first helical turns of S3 and S4 but not with similarly located Cys in S1 or S2. Thus, in the membrane, S0 is next to S3 and S4 but not to S1 and S2, although the flank of S0 reaches the flanks of S1 and S2. Furthermore, co-expression of the double-Cys mutant of alpha, W23C in the first helical turn of S0 and F144C in the S2 flank, and the double-Cys mutant of beta1, Y42C in the TM1 flank and L157C in the first helical turn of TM2, resulted in the crosslinking of two alphas through one beta1, S0 to TM2 and TM1 to S2. Thus, TM1 and TM2 of each beta subunit lie between the voltage-sensing domains (S0-S4) of adjacent alpha subunits.