Transfer of L-type calcium channel IVS6 segment increases phenylalkylamine sensitivity of α1A

Abstract

Conditioned ("use-dependent") inhibition by phenylalkylamines (PAAs) is a characteristic property of L-type calcium (Ca2+) channels. To determine the structural elements of the PAA binding domain we transferred sequence stretches of the pore-forming regions of repeat III and/or IV from the skeletal muscle αt subunit (α1s) to the class A α1 subunit (α1A) and expressed these chimeras together with 1a and α2/δ subunits in Xenopus oocytes. The corresponding barium currents (IBa) were tested for PAA sensitivity during trains of depolarizing test pulses (conditioned block). IBa of oocytes expressing the α1A subunit were only weakly inhibited by PAAs (less than 10% conditioned block of IBa during a 100-ms pulse train of 0.1 Hz). Transfer of the transmembrane segment IVS6 from α1s to α1A produced an enhancement of PAA sensitivity of the resulting α1A/α1s chimera comparable to L-type α1 subunits (about 35% conditioned block of IBa during a 100-ms pulse train of 0.1 Hz). Our results demonstrate that substitution of 11 amino acids within the segment IVS6 of α1A with the corresponding residues of α1s is sufficient to transfer L-type PAA sensitivity into the low sensitive class A Ca2+ channel.