Voltage Gated Potassium Channel Assays
Voltage gated potassium channels are potassium-specific transmembrane channels that respond to voltage changes in the cell's membrane potential. They are important in restoring the depolarized cell to a resting state during action potentials.
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Structure
Voltage gated potassium channels in vertebrates are tetramers made up of four identical subunits organized in a ring, each contributing to the trans-membrane K+ pore wall. Each subunit is made up of six membrane-spanning hydrophobic -helical sequences and a voltage sensor. Both amino and carboxy termini can be found on the intracellular side of the membrane.
Feature
Voltage gated potassium channels are selective for K+ over other cations like Na+. At the narrowest section of the transmembrane pore, there is a selectivity filter. Water molecules solvate potassium and sodium cations in an aqueous environment. When the potassium channel passes through its selectivity filter, the water-K+ interactions are replaced by interactions between K+ and the channel protein's carbonyl groups. The selectivity filter's diameter is suitable for potassium cations but too large for the smaller sodium cations. As a result, the protein carbonyl groups effectively "solvate" the potassium cations, but these same carbonyl groups are too widely apart to adequately solvate the sodium cation.
Published Data
| Paper Title | Intracellular hemin is a potent inhibitor of the voltage-gated potassium channel Kv10.1 |
| Journal | Scientific Reports |
| Published | 2022 |
| Abstract | Heme, a cofactor that is attached to different hemoproteins, promotes a wide range of processes, including signal transduction, oxygen transport, electron transfer, and drug metabolism. Here, they demonstrate how human voltage-gated potassium channels (hEAG1, Kv10.1) are modified by intracellular free heme and hemin. Hemin applied to the intracellular side potently inhibits Kv10.1 channels, favoring inhibition at resting potential, with an IC50 of around 4 nM under ambient circumstances and 63 nM under lowering conditions. |
| Result |
They expressed the channel-coding mRNA in Xenopus oocytes and used the inside-out mode of the patch-clamp technique to record voltage-dependent currents in membrane patches big enough to produce macroscopic currents in order to determine how sensitive Kv10.1 channels are to intracellular hemin or heme. A. At the test voltages applied, the current amplitudes were gradually reduced by increasing hemin concentrations, with 10 nM suppressing the current by more than 50%. B. Following the administration of hemin, the current decreased exponentially, achieving an almost total loss of current at 100 nM in about a minute. C. The reducing agent DTT was used to hasten the inhibitory effect of hemin. D. With 1 mM GSH present, the IC50 increased from around 4 nM to 63 nM. Cysteine residues may be involved given that the effect of hemin on Kv10.1 channels is redox dependent.
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Fig.1. Kv10.1 current inhibition by hemin. (Wall, 2022)
Reference
- Sahoo, N.; et al. Intracellular hemin is a potent inhibitor of the voltage-gated potassium channel Kv10.1. Scientific reports. 2022, 12(1): 1-12.
