Detection of nucleotide binding to Na,K-ATPase in an aqueous membrane suspension by 13C cross-polarization magic-angle spinning NMR spectroscopy.
Binding of uniformly (13)C labelled ATP to Na,K-ATPase was studied by (13)C cross-polarization magic-angle spinning (CP-MAS) NMR. In the presence of 30 mM Na(+) , and with sample- and time-averaging, NMR spectra obtained at 4 degrees C exhibited several resonances for the bound nucleotide. Chemical shifts suggested that site-specific changes in the micro-environment or conformation of the nucleotide occurred in the high affinity binding site. These experiments permit further studies of nucleotide dynamics, structure and binding under physiologically relevant conditions.
Middleton
David A
DA
School of Biological Sciences, University of Liverpool, Crown Street, Liverpool L79 7ZB, United Kingdom.
Jakobsen
Louise Odgaard
LO
Esmann
Mikael
M
D000818:Animals; D002247:Carbon Isotopes; D006868:Hydrolysis; D007668:Kidney / Q000201:enzymology; D008566:Membranes / Q000201:enzymology; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D009711:Nucleotides / Q000737:chemistry / Q000378:metabolism*; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D013552:Swine
D002247:Carbon Isotopes; D009711:Nucleotides; D000254:Sodium-Potassium-Exchanging ATPase