An artificial tetramerization domain restores efficient assembly of functional Shaker channels lacking T1

One feature shared by all Shaker-type voltage-gated K+ channels is a highly conserved domain (T1) located in the cytoplasmic N terminus. The T1 domain is a key determinant of which subtypes can form heteromultimeric channels, suggesting that T1 functions during channel assembly. To better define the role of T1 during channel assembly and separate this function from potenti al contributions to channel permeation and gating, we replaced the T1 domai n (residues 96-183) of ShakerB with a coiled-coil sequence (GCN4-LI) that f orms parallel tetramers. Deleting T1 dramatically, but not completely. abol ished channel formation under most expression conditions. Channels lacking T1 are functional and K+-selective, although they activate at more hyperpol arized membrane potentials and inactivate less completely. Insertion of the artificial tetramerization domain (GCN4-LI) restored efficient channel for mation, suggesting that tetramerization of the cytoplasmic T1 domain promot es transmembrane channel assembly by increasing the effective local subunit concentration for T1 compatible subunits, We propose that T1 tetramerizati on promotes subfamily-specific assembly through kinetic partitioning of the assembly process, but is not required for subsequent steps in channel asse mbly and folding.