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Labeling of human erythrocyte membranes with eosin probes used for protein diffusion measurements : inhibition of anion transport and photo-oxidative inactivation of acetylcholinesterase

Nigg, E. and Kessler, M. and Cherry, R. J.. (1979) Labeling of human erythrocyte membranes with eosin probes used for protein diffusion measurements : inhibition of anion transport and photo-oxidative inactivation of acetylcholinesterase. Biochimica et biophysica acta, Vol. 550, H. 2. S. 328-340.

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Official URL: http://edoc.unibas.ch/dok/A5249535

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Abstract

The binding of eosin-isothiocyanate (eosin-NCS) and iodoacetamido-eosin (IA-eosin) to band 3 proteins in the membrane of human erythrocytes is characterized by studying the effect of these probes on the anion transport system. Although the unbrominated fluorescein precursors do not affect anion transport, both eosin labels are strong inhibitors of sulphate exchange in intact erythrocytes. 50% inhibition is obtained by binding 4.7 . 10(5) or 6.0 . 10(5) molecules/cell for eosin-NCS and IA-eosin, respectively. Both eosin probes are irreversibly bound and occupy common binding sites with 4,4-diisothiocyano-1,2-diphenyl-ethane-2,2'disulfonic acid (H2DIDS), although other sites are labeled as well. The inhibition of anion transport is light independent and can therefore not be attributed to a photosensitizing action of the eosin probes. Both eosin derivatives, however, inactivate acetylcholinesterase upon illumination of air-equilibrated samples of hemoglobin-free labeled ghosts. The inactivation of the enzyme is accompanied by the formation of protein aggregates as visualized by polyacrylamide gel electrophoresis. These effects are not observed when intact erythrocytes are illuminated in the presence of eosin probes suggesting a protective effect of hemoglobin during the labeling procedure. Protection of ghosts from photo-oxidation is achieved by displacing air with argon. These results are discussed in relation to the use of these and similar probes to measure protein diffusion in membranes.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum
05 Faculty of Science > Departement Biozentrum > Growth & Development > Cell Biology (Nigg)
UniBasel Contributors:Nigg, Erich A.
Item Type:Article, refereed
Bibsysno:Link to catalogue
Publisher:Elsevier
ISSN:0006-3002
Note:Publication type according to Uni Basel Research Database: Journal article
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Last Modified:22 Mar 2012 14:19
Deposited On:22 Mar 2012 13:17

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