Calcium entry through voltage-gated calcium channels (VGCC) initiates diverse cellular functions. VGCC pore-forming subunit (CaVα 1) contains four homology repeats, each encompassing a voltage sensor and a pore domain. Three main classes of CaVα1 subunits have been described, CaV1, CaV2 and Ca V3 that differ in their voltage-dependence of activation and in the extent in which this process is modulated by the auxiliary β-subunit (CaVβ). Association of CaVβ induces a coil-to-helix conformation of the I-II intracellular linker joining the first and second repeat of CaVα1 that is thought to be crucial for modulation of channel function. When expressed in Xenopus laevis oocytes in the absence of CaVβ, the voltage to reach 50% activation (V 0.5) for CaV1.2 and CaV2.3 differs by more than 60 mV and the channel current-carrying capacity by more than thirty-fold. Here we report that the difference in V0.5 is reduced to about 30 mV and the current-carrying capacity becomes virtually identical when the I-II linkers of CaV1.2 and CaV2.3 are swapped. Co-expression with CaVβ increases the current-carrying capacity of chimeric channels by the same extent, while the difference in V0.5 with respect to their corresponding parental channels vanishes. Our findings indicate that CaVβ modulatory potency is determined by both, the nature of the I-II linker and the pore-forming subunit background. Moreover, they demonstrate that the I-II linker encodes self-reliant molecular determinants for channel activation and suggest that besides the secondary structure adopted by this segment upon CaVβ association, its chemical nature is as well relevant. © 2010 Landes Bioscience.