ATP-binding is stabilized by a stacking interaction within the binding site of Na+/K+ -ATPase.
Site-directed mutagenesis was applied to modify phenylalanines (Phe(475)Trp, Phe(548)Tyr, and both) to generate mutants on the basis of molecular modeling of the ATP-binding domain of Na(+)/K(+)-ATPase, in order to characterize the forces that stabilize ATP in its binding pocket. Each of the mutants was examined by Raman difference spectroscopy, i.e., as a difference between the spectrum of the domain with and without bound ATP. It was shown that Phe(475) plays a key role in stabilizing ATP-binding by a stacking interaction. Phe(548) co-stabilizes ATP on the opposite site of the binding pocket and its type of interaction with ATP-binding differs from that of Phe(475).
Hofbauerová
Katerina
K
Institute of Physiology, Academy of Sciences of the Czech Republic, Vídenská 1083, CZ-14220 4, Prague, Czech Republic.
Kopecký
Vladimír
V
Ettrich
Rüdiger
R
Kubala
Martin
M
Teisinger
Jan
J
Amler
Evzen
E
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D051379:Mice; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010649:Phenylalanine / Q000737:chemistry; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry* / Q000378:metabolism; D013059:Spectrum Analysis, Raman
D010649:Phenylalanine; D000255:Adenosine Triphosphate; D000254:Sodium-Potassium-Exchanging ATPase