title: Polarized infrared absorption of Na+/K+-ATPase studied by attenuated total reflection spectroscopy.
abstract: Na+/K+-ATPase can be isolated from the outer medulla of mammalian kidney in the form of flat membrane fragments containing the enzyme in a density of 10(3)-10(4) protein molecules per microm2 (Deguchi et al. (1977) J. Cell. Biol. 75, 619-634). In this paper we show that these membrane fragments can be bound to a germanium plate coated with a phospholipid bilayer. With this system infrared spectroscopic studies of the enzyme have been carried out using the technique of attenuated total reflection (ATR). At a coverage of the lipid surface corresponding to 30-40% of a monolayer of membrane fragments, characteristic infrared bands of the protein such as the amide I and II bands can be resolved. About 24% of the NH-groups of the peptide backbone are found to be resistant to proton/deuterium exchange within a time period of several days. Evidence for orientation of the protein with respect to the supporting lipid layer is obtained from experiments with polarized light, the largest polarization effects being associated with the -COO- band at 1400 cm-1. Experiments with aqueous media of different ionic composition indicate that the average orientation of transition moments changes when K+ in the medium is replaced by Tris+ or Na+.
authors.affiliation: Department of Biology, University of Konstanz, F.R.G.
authors.forename: U P
authors.initials: UP
authors.lastname: Fringeli
authors.forename: H J
authors.initials: HJ
authors.lastname: Apell
authors.forename: M
authors.initials: M
authors.lastname: Fringeli
authors.forename: P
authors.initials: P
authors.lastname: Läuger
mesh_terms: D000818:Animals; D002021:Buffers; D002414:Cations, Monovalent; D002462:Cell Membrane / Q000648:ultrastructure; D066298:In Vitro Techniques; D007679:Kidney Medulla / Q000201:enzymology; D008563:Membrane Lipids; D008565:Membrane Proteins; D011487:Protein Conformation; D011817:Rabbits; D000254:Sodium-Potassium-Exchanging ATPase; D013055:Spectrophotometry, Infrared / Q000379:methods
chemical_list: D002021:Buffers; D002414:Cations, Monovalent; D008563:Membrane Lipids; D008565:Membrane Proteins; D000254:Sodium-Potassium-Exchanging ATPase