The function of BK channels extracted and purified within SMALPs

Human BK channels are large voltage and Ca2+-activated K+ channels, involved in several important functions within the body. The core channel is a tetramer of alpha subunits, and its function is modulated by the presence of beta and gamma accessory subunits. BK channels composed of alpha subunits, as well as BK channels composed of alpha and beta 1 subunits, were successfully solubilised from HEK cells with styrene maleic acid (SMA) polymer and purified by nickel affinity chromatography. Native SMA-PAGE analysis of the purified proteins showed the alpha subunits were extracted as a tetramer. In the presence of beta 1 subunits, they were co-extracted with the alpha subunits as a heteromeric complex. Purified SMA lipid particles (SMALPs) containing BK channel could be inserted into planar lipid bilayers (PLB) and single channel currents recorded, showing a high conductance (approximate to 260 pS), as expected. The open probability was increased in the presence of co-purified beta 1 subunits. However, voltage-dependent gating of the channel was restricted. In conclusion, we have demonstrated that SMA can be used to effectively extract and purify large, complex, human ion channels, from low expressing sources. That these large channels can be incorporated into PLB from SMALPs and display voltage-dependent channel activity. However, the SMA appears to reduce the voltage dependent gating of the channels.