title: Continuous fluorescence-based measurement of redox-driven sodium ion translocation.
abstract: Investigation of the mechanism of sodium ion pumping enzymes requires methods to follow the translocation of sodium ions by the purified and reconstituted proteins in vitro. Here, we describe a protocol that allows following the accumulation of Na(+) in proteoliposomes by the Na(+)-translocating NADH:quinone oxidoreductase (Na(+)-NQR) from Vibrio cholerae using the sodium-sensitive fluorophor sodium green. In the presence of a regenerative system for its substrate NADH, the Na(+)-NQR accumulates Na(+) in the proteoliposomes which is visible as a change in fluorescence.
authors.affiliation: Department of Microbiology, University of Hohenheim (Stuttgart), Garbenstrasse 30, 70599 Stuttgart, Germany.
authors.forename: Valentin
authors.initials: V
authors.lastname: Muras
authors.affiliation: Department of Microbiology, University of Hohenheim (Stuttgart), Garbenstrasse 30, 70599 Stuttgart, Germany.
authors.forename: Björn
authors.initials: B
authors.lastname: Claussen
authors.affiliation: European Molecular Biology Laboratory (EMBL), Grenoble Outstation, 6 rue Jules Horowitz, 38042 Grenoble, France; Unit for Virus Host-Cell Interactions, University Grenoble Alpes-EMBL-CNRS, 6 rue Jules Horowitz, 38042 Grenoble, France.
authors.forename: Manikandan
authors.initials: M
authors.lastname: Karuppasamy
authors.affiliation: European Molecular Biology Laboratory (EMBL), Grenoble Outstation, 6 rue Jules Horowitz, 38042 Grenoble, France; Unit for Virus Host-Cell Interactions, University Grenoble Alpes-EMBL-CNRS, 6 rue Jules Horowitz, 38042 Grenoble, France.
authors.forename: Christiane
authors.initials: C
authors.lastname: Schaffitzel
authors.affiliation: Department of Microbiology, University of Hohenheim (Stuttgart), Garbenstrasse 30, 70599 Stuttgart, Germany. Electronic address: julia.steuber@uni-hohenheim.de.
authors.forename: Julia
authors.initials: J
authors.lastname: Steuber
mesh_terms: D001692:Biological Transport; D015374:Biosensing Techniques / Q000379:methods*; D009243:NAD / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D011808:Quinone Reductases / Q000378:metabolism; D012964:Sodium / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D014734:Vibrio cholerae / Q000201:enzymology
chemical_list: D009243:NAD; D012964:Sodium; C041831:NADH dehydrogenase (quinone); D011808:Quinone Reductases
keywords: Fluorescence; Proteoliposomes; Reconstitution; Sodium green; Sodium ion-translocating NADH:quinone oxidoreductase (Na(+)-NQR); Sodium transport