Significance β-Subunits (β1–β4) play a critical role in defining the properties of the voltage- and calcium-activated potassium (BK) channel, which in turn determines the physiological role that this channel can perform in different tissues. In particular, the β1-subunit causes an increase in the apparent BK Ca 2+ sensitivity due to a stabilization of the voltage sensor in the active configuration. We investigated the molecular details of such voltage-sensor stabilization by mutagenesis and gating current measurements. Mixing regions of β1 and β3 made it possible to identify the N terminus, in particular the third and fourth lysine residues, as the structural element necessary to recover the full effect of β1 on the voltage sensor.