Engineered Passive Potassium Conductance in the KR2 Sodium Pump
| Autor: | Christiane Grimm, Arend Vogt, Maximiliano Ancina Möller, Peter Hegemann, Florian Heiser, Arita Silapetere |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
Light
Protein Conformation Potassium Sodium Biophysics chemistry.chemical_element Molecular engineering Ion Cell Line Cell membrane 03 medical and health sciences Mice 0302 clinical medicine Rhodopsins Microbial medicine Escherichia coli Animals Ion transporter 030304 developmental biology chemistry.chemical_classification 0303 health sciences Ion Transport Cell Membrane Conductance Biological Transport Articles Hydrogen-Ion Concentration Rats medicine.anatomical_structure chemistry Mutant Proteins Counterion Sodium-Potassium-Exchanging ATPase Flavobacteriaceae 030217 neurology & neurosurgery Protein Binding |
| Zdroj: | Biophys J |
| Popis: | Light-driven sodium pumps (NaRs) are microbial rhodopsins that utilize light energy to actively transport sodium ions out of the cell. Here, we used targeted mutagenesis and electrophysiological methods in living cells to demonstrate that NaRs can be converted into light-activated cation channels by molecular engineering. Specifically, introduction of the R109Q mutation into the sodium ion pump of Dokdonia eikasta (KR2) results in passive ion conductance, with a high preference for potassium over sodium ions. However, in this mutant, residual active outward pumping of sodium ions competes with passive inward transport of potassium. Channel-like behavior could also be achieved by introduction of other mutations into the KR2 counterion complex, and further, these modifications were transferrable to other NaRs. Combining the R109Q replacement with modifications at position S70 removed the residual sodium pumping and greatly enhanced the channel-like activity. However, passive photocurrents were only observed in leak mutants if the KR2 counterions, D116 and D251, were deprotonated, which was only observed under alkaline conditions. Overall, our results reveal that interactions between R109 and the nearby residues, L75, S70, D116, and D251, prevent passive backflow during ion transport in NaRs. |
| Databáze: | OpenAIRE |
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