Electron spin resonance studies on the inorganic-anion-transport system of the human red blood cell Binding of a disulfonatostilbene spin label (NDS-tempo) and inhibition of anion transport

Schnell, K. F.; Elbe, W.; Käsbauer, J; Kaufmann, E.

doi:10.1016/0005-2736(83)90211-0

Cited by 18 publications

(12 citation statements)

References 31 publications

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“…) which bind preferentially to Band 3 and which are irreversible inhibitors of the anion transport in red blood cells, completely prevent the NDS-TEMPO binding to the erythrocyte membrane. Third, the addition of DIDS or of DNDS to NDS-TEMPOlabeled red blood cells results in an immediate release of bound NDS-TEMPO from the membrane surface, a disappearance of the immobile signal and a concomitant increase of the mobile ESR signal (Schnell et al, 1983). ...... Membrane-bound NDS-TEMPO appear~:~m be almost completely immobilized.…”

Section: Discussionmentioning

confidence: 97%

“…Preceding studies have shown that NDS-TEMPO is a nonpenetrating, competitive inhibitor of the chloride and the sulfate transport in human red cell ghosts. Concomitantly, chloride, sulfate and other substrate anions are competitive inhibitors of NDS-TEMPO binding to the erythrocyte membrane (Schnell et al, 1981c(Schnell et al, , 1983. The mutual competition between NDS-TEMPO binding and substrate-anion binding provides strong evidence for the assumption that NDS-TEMPO in fact binds to the substrate site of Band 3.…”

Section: Introductionmentioning

confidence: 93%

“…Because of the high dielectric loss caused by the aqueous solutions, a high microwave power was required for the measurements. Care was taken to avoid distortions of the ESR spectra which could result from saturation effects or inappropriate choice of the scan time and of the time constant (Jost & Griffith, 1976;Schnell et al, 1983). The temperature of the samples was controlled and amounted to 20~ during microwave irradiation.…”

Section: Esr Spectramentioning

confidence: 99%

“…Mol wt: 689; purity 96% (HPLC), water soluble. For synthesis and purification see Schnell et al (1983) & Passow, 1979;Rao et al, 1979;Fr6hlich, 1982;Furuya et al, 1984). We therefore have synthesized the disulfonatostilbene spin label NDS-TEMPO.…”

Section: Introductionmentioning

confidence: 99%

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Characterization of the Band 3 substrate site in human red cell ghosts by NDS-TEMPO, a disulfonatostilbene spin probe: The function of protons in NDS-TEMPO and substrate-anion binding in relation to anion transport

1986

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NDS-TEMPO is a specific disulfonatostilbene spin label for the Band 3 substrate site (K.F. Schnell, W. Elbe, J. Käsbauer & E. Kaufmann, Biochim. Biophys. Acta 732:266-275, 1983). The pH dependence of NDS-TEMPO binding and of chloride and sulfate binding was studied in resealed human erythrocyte ghosts. pH was varied from 6.0 to 9.0. The ESR spectra from NDS-TEMPO-labeled red cell ghosts exhibited a strong immobilization of membrane-bound NDS-TEMPO. Changes of pH had no effect upon the mobility of membrane-bound NDS-TEMPO. A mutual competition between NDS-TEMPO binding and the binding of the substrate-anions, chloride and sulfate, was observed throughout the entire pH range. The maximal number of NDS-TEMPO binding sites per cell was in the range of 9.0 X 10(5) to 1.10 X 10(6) and was found to be insusceptible to changes of pH. The NDS-TEMPO/substrate-site and the chloride/substrate-site dissociation constants amounted to 1.25 microM and to 17 mM and were independent of pH from pH 6.0 to 8.0, while the sulfate/substrate-site dissociation constant displayed a strong pH dependency with a maximum of approximately 50 mM at about pH 7.0. The NDS-TEMPO inhibition constants from the chloride and the sulfate flux experiments were 0.5 microM (0 degree C) and 1.8 microM (25 degrees C), respectively, and are in close accordance with the NDS-TEMPO/substrate-site dissociation constants. Our studies provide strong evidence for the assumption that NDS-TEMPO binds in fact to the substrate site of Band 3. They show that the strong pH dependence of the chloride and of the sulfate transport cannot result from the pH dependency of substrate-anion binding, but point to the participation of ionizable regulator sites in transport catalysis. These regulator sites seem to be positioned outside the substrate site of the Band 3 transport domain.

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Section: Discussionmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 93%

Section: Esr Spectramentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Characterization of the Band 3 substrate site in human red cell ghosts by NDS-TEMPO, a disulfonatostilbene spin probe: The function of protons in NDS-TEMPO and substrate-anion binding in relation to anion transport

1986

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“…Fr6hlich [38] has shown that DNDS binds to a single class of sites, indicating that binding of DNDS to one band 3 protomer does not affect the DNDS affinity for the other subunit(s) of a dimer or tetramer. Schnell et al [126] synthesized a spin-labeled stilbenedisulfonate, NDS-TEMPO, one end of which is rather bulky. Both in the presence and absence of transportable anions, the binding of NDS-TEMPO to the membrane is a simple hyperbolic function of the free concentration over the low-to-moderate concentration range.…”

Section: Reversible Stilbenedisulfonate Bindingmentioning

confidence: 99%

Oligomeric structure and the anion transport function of human erythrocyte band 3 protein

Jennings

1984

J. Membrain Biol.

149

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Molecular aspects of band 3 protein-mediated anion transport across the red blood cell membrane

Passow

1986

Reviews of Physiology, Biochemistry and Pharmacology, Volume 103

157

139

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Electron spin resonance studies on the inorganic-anion-transport system of the human red blood cell Binding of a disulfonatostilbene spin label (NDS-tempo) and inhibition of anion transport

Cited by 18 publications

References 31 publications

Characterization of the Band 3 substrate site in human red cell ghosts by NDS-TEMPO, a disulfonatostilbene spin probe: The function of protons in NDS-TEMPO and substrate-anion binding in relation to anion transport

Characterization of the Band 3 substrate site in human red cell ghosts by NDS-TEMPO, a disulfonatostilbene spin probe: The function of protons in NDS-TEMPO and substrate-anion binding in relation to anion transport

Oligomeric structure and the anion transport function of human erythrocyte band 3 protein

Molecular aspects of band 3 protein-mediated anion transport across the red blood cell membrane

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