Molecular rearrangements of the Kv2.1 potassium channel termini associated with voltage gating

The voltage-gated K(v)2.1 channel is composed of four identical subunits folded around the central pore and does not inactivate appreciably during short depolarizing pulses. To study voltage-induced relative molecular rearrangements of the channel, K(v)2.1 subunits were genetically fused with enhanced cyan fluorescent protein and/or enhanced yellow fluorescent protein, expressed in COS1 cells, and investigated using fluorescence resonance energy transfer ( FRET) microscopy combined with patch clamp. Fusion of fluorophores to either or both termini of the K(v)2.1 monomer did not significantly affect the gating properties of the channel. FRET between the N- and C-terminal tags fused to the same or different K(v)2.1 monomers decreased upon activation of the channel by depolarization from - 80 to + 60 mV, suggesting voltage-gated relative rearrangement between the termini. Because FRET between the K(v)2.1 N- or C-terminal tags and the membrane-trapped EYFPN-PH pleckstrin homology domains did not change on depolarization, voltage-gated relative movements between the K(v)2.1 termini occurred in a plane parallel to the plasma membrane, within a distance of 1-10 nm. FRET between the N-terminal tags did not change upon depolarization, indicating that the N termini do not rearrange relative to each other, but they could either move cooperatively with the K(v)2.1 tetramer or not move at all. No FRET was detected between the C-terminal tags. Assuming their randomized orientation in the symmetrically arranged K(v)2.1 subunits, C termini may move outwards in order to produce relative rearrangements between N and C termini upon depolarization.