Charge translocation by the Na,K-pump: II. Ion binding and release at the extracellular face
| Autor: | R. Bühler, Hans-Jürgen Apell, Peter Läuger, W. Stürmer |
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| Rok vydání: | 1991 |
| Předmět: |
Physiology
Stereochemistry Tetraphenylborate Biophysics Biological Transport Active Diaphragm pump In Vitro Techniques chemistry.chemical_compound Adenosine Triphosphate Onium Compounds Organophosphorus Compounds Ion binding Extracellular Animals Phosphorylation Na+/K+-ATPase Lipid bilayer Binding Sites Cell Membrane Osmolar Concentration Sodium Cell Biology Electrophysiology Spectrometry Fluorescence Membrane chemistry Ionic strength Potassium Sodium-Potassium-Exchanging ATPase Extracellular Space |
| Zdroj: | The Journal of Membrane Biology. 121:163-176 |
| ISSN: | 1432-1424 0022-2631 |
| DOI: | 10.1007/bf01870530 |
| Popis: | In the first part of the paper, evidence has been presented that electrochromic styryl dyes, such as RH 421, incorporate into Na,K-ATPase membranes isolated from mammalian kidney and respond to changes of local electric field strength. In this second part of the paper, fluorescence studies with RH-421-labeled membranes are described, which were carried out to obtain information on the nature of charge-translocating reaction steps in the pumping cycle. Experiments with normal and chymotrypsin-modified membranes show that phosphorylation by ATP and occlusion of Na+ are electroneutral steps, and that release of Na+ from the occluded state to the extracellular side is associated with translocation of charge. Fluorescence signals observed in the presence of K+ indicate that binding and occlusion of K+ at the extracellular face of the pump is another major electrogenic reaction step. The finding that the fluorescence signals are insensitive to changes of ionic strength leads to the conclusion that the binding pocket accommodating Na+ or K+ is buried in the membrane dielectric. This corresponds to the notion that the binding sites are connected with the extracellular medium by a narrow access channel ("ion well"). This notion is further supported by experiments with lipophilic ions, such as tetraphenylphosphonium (TPP+) or tetraphenylborate (TPB-), which are known to bind to lipid bilayers and to change the electrostatic potential inside the membrane. Addition of TPP+ leads to a decrease of binding affinity for Na+ and K+, which is thought to result from the TPP(+)-induced change of electric field strength in the access channel. |
| Databáze: | OpenAIRE |
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