Site-specific ion occupation in the selectivity filter causes voltage-dependent gating in a viral K+ channel

Abstract

Many potassium channels show voltage-dependent gating without a dedicated voltage sensor domain. This is not fully understood yet, but often explained by voltage-induced changes of ion occupation in the five distinct K+ binding sites in the selectivity filter. To better understand this mechanism of filter gating we measured the single-channel current and the rate constant of sub-millisecond channel closure of the viral K+ channel KcvNTS for a wide range of voltages and symmetric and asymmetric K+ concentrations in planar lipid membranes. A model-based analysis employed a global fit of all experimental data, i.e., using a common set of parameters for current and channel closure under all conditions. Three different established models of ion permeation and various relationships between ion occupation and gating were tested. Only one of the models described the data adequately. It revealed that the most extracellular binding site (S0) in the selectivity filter functions as the voltage sensor for the rate constant of channel closure. The ion occupation outside of S0 modulates its dependence on K+ concentration. The analysis uncovers an important role of changes in protein flexibility in mediating the effect from the sensor to the gate.

Unravelling the intricate cooperativity of subunit gating in P2X2 ion channels

“…Global fits provided also essentially new insights into the function of other ion channels, e.g. for the gating charge movement in Shaker channels 21 and for the effects of permeating K + ions on the IV -relationships of viral Kv channels 20 , all measured in ensemble currents and under equilibrium conditions. Also single-channel currents were subjected to global fit approaches, as shown elegantly e.g.…”

“…Major progress in understanding the function of P2X receptors can be expected from applying global fit analyses of extended data, as demonstrated previously for other ion channels. Using current data at equilibrium this has been demonstrated for concentration-activation relationships of concatenated CNGA2 channels, containing a different number of disabled binding sites 19 , for IV -relationships of viral Kv channels at different concentrations of permeating K + ions 20 , or gating charge movement in Shaker channels 21 . We previously also globally fitted data distant from equilibrium by kinetic schemes, for activation time courses of tetrameric CNGA2 channels induced by cGMP jumps to different concentrations 22 and voltage-induced activation time courses of tetrameric HCN2 channels 23 .…”

Photolithographic Fabrication of Micro Apertures in Dry Film Polymer Sheets for Channel Recordings in Planar Lipid Bilayers

“…In subsequent experiments, we compared the functional properties of a reference K + channel, the small viral protein Kcv NTS (Rauh et al 2018 ), that has been reconstituted in lipid bilayers formed over ADEX- and Teflon-based septa. Representative recordings of single channel fluctuations in a DPhPC bilayer generated over an approx.…”

“…At negative voltages of approx. − 100 mV, the unitary openings become increasingly noisy; the latter is caused by a typical fast gating at negative voltages, which cannot be resolved in conventional recording set-up (Rauh et al 2018 ). From the unitary channel fluctuations, we constructed the current/voltage relation as well as the open probability/voltage in both recording conditions (Fig.…”

A Functional K+ Channel from Tetraselmis Virus 1, a Member of the Mimiviridae

“…This includes the same mechanism of ion selectivity, which provides the channel with a distinct preference for transport of K + over Na + as well as a sensitivity to Ba 2+ block [ 7 , 9 , 12 ]. Moreover, like their prokaryotic and eukaryotic relatives, the viral channel is gated, meaning that it switches stochastically between defined open and closed states [ 12 , 13 ].…”