The miniature viral K+ channel Kcv represents the pore module of all K+ channels. A synthetic
gene of Kcv with an elevated GC content compared to that of the wild-type gene was expressed
heterologously in Pichia pastoris, and the purified protein was functionally reconstituted into liposomes.
Biochemical assays reveal a remarkable cation selctive stability of the channel tetramer via SDS−PAGE.
Only cations, which permeate Kcv, were able to protect the oligomer against disassembly into monomers
at high temperatures. Electrophysiological characterization of the single Kcv channel reveals a saturating
conductance (Λmax) of 360 pS; the single-channel current−voltage relation was strongly rectifying with
a negative slope conductance at extreme voltages. The channel was highly selective for K+ and was
blocked by Ba2+ and in a side specific manner by Na+ and Cs+ also. The channel conducted Rb+, but as
a consequence, the channel was shifted into a hyperactive state. We conclude that specific binding
interactions of cations in the conductive pathway are an important determinant of channel stability and
function.