Disulfide Crosslinking Between BK Channel Alpha And Beta1 Subunits In The Membrane Domain

Abstract

The large-conductance, Ca2+- and voltage-activated potassium channel (BK) alpha subunit is modulated by one of four types of beta subunits, each imparting unique electrophysiological properties. The beta1 subunit is expressed in smooth muscle, where it renders the BK channel sensitive to cytoplasmic calcium in a voltage range near the smooth-muscle resting potential and slows activation and deactivation. Previously, we inferred from the extents of endogenous disulfide bond formation between cysteines substituted in the extracellular flanks of the transmembrane (TM) helices of alpha and of beta1, that the extracellular flank of S0, the unique seventh TM helix of BK alpha, is surrounded on three sides by the extracellular flanks of S1 and S2 and the four-residue, extracellular loop between S3 and S4. We also found that the extracellular flanks of beta1 TM2 and alpha S0 are close and that that the extracellular end of beta1 TM1 is close to the extracellular flanks of both alpha S1 and S2. Within the membrane domain, where presumably there is less flexibility than in the extracellular flanks, we also see endogenous (no added reagents) crosslinking one to two helical turns into the membrane domain. Within the same alpha subunit, Cys-substituted residues in S0 readily form disulfides with Cys-substituted residues in S4, and to a lesser extent with Cys-substituted residues in S3. Also within the membrane domain, we find that cysteines in alpha S0 readily form disulfides with cysteines in beta TM2. Thus, the positions of alpha S0 and of beta1 TM2 relative to alpha S0 are similar in the membrane domain and in the extracellular flanks of these TM helices.Supported by HL081172 and NS054946