Competition between Na(+) and Li(+) for unsealed and cytoskeleton-depleted human red blood cell membrane: a (23)Na multiple quantum filtered and (7)Li NMR relaxation study.
Evidence for competition between Li(+) and Na(+) for binding sites of human unsealed and cytoskeleton-depleted human red blood cell (csdRBC) membranes was obtained from the effect of added Li(+) upon the (23)Na double quantum filtered (DQF) and triple quantum filtered (TQF) NMR signals of Na(+)-containing red blood cell (RBC) membrane suspensions. We found that, at low ionic strength, the observed quenching effect of Li(+) on the (23)Na TQF and DQF signal intensity probed Li(+)/Na(+) competition for isotropic binding sites only. Membrane cytoskeleton depletion significantly decreased the isotropic signal intensity, strongly affecting the binding of Na(+) to isotropic membrane sites, but had no effect on Li(+)/Na(+) competition for those sites. Through the observed (23)Na DQF NMR spectra, which allow probing of both isotropic and anisotropic Na(+) motion, we found anisotropic membrane binding sites for Na(+) when the total ionic strength was higher than 40 mM. This is a consequence of ionic strength effects on the conformation of the cytoskeleton, in particular on the dimer-tetramer equilibrium of spectrin. The determinant involvement of the cytoskeleton in the anisotropy of Na(+) motion at the membrane surface was demonstrated by the isotropy of the DQF spectra of csdRBC membranes even at high ionic strength. Li(+) addition initially quenched the isotropic signal the most, indicating preferential Li(+)/Na(+) competition for the isotropic membrane sites. High ionic strength also increased the intensity of the anisotropic signal, due to its effect on the restructuring of the membrane cytoskeleton. Further Li(+) addition competed with Na(+) for those sites, quenching the anisotropic signal. (7)Li T(1) relaxation data for Li(+)-containing suspensions of unsealed and csdRBC membranes, in the absence and presence of Na(+) at low ionic strength, showed that cytoskeleton depletion does not affect the affinity of Na(+) for the RBC membrane, but increases the affinity of Li(+) by 50%. This clearly indicates that cytoskeleton depletion favors Li(+) relative to Na(+) binding, and thus Li(+)/Na(+) competition for its isotropic sites. Thus, this relaxation technique proves to be very sensitive to alkali metal binding to the membrane, detecting a more pronounced steric hindrance effect of the cytoskeleton network to binding of the larger hydrated Li(+) ion to the membrane phosphate groups.
Srinivasan
C
C
Department of Chemistry, Loyola University of Chicago, 6525 N. Sheridan Road, Chicago, Illinois 60626, USA.
Minadeo
N
N
Toon
J
J
Graham
D
D
Mota de Freitas
D
D
Geraldes
C F
CF
D000465:Algorithms; D001667:Binding, Competitive; D016009:Chi-Square Distribution; D004910:Erythrocyte Membrane / Q000378:metabolism*; D005583:Fourier Analysis; D006801:Humans; D016018:Least-Squares Analysis; D008094:Lithium / Q000097:blood*; D009682:Magnetic Resonance Spectroscopy; D012964:Sodium / Q000097:blood*
D008094:Lithium; D012964:Sodium