Interaction of calmodulin with the red cell and its membrane skeleton and with spectrin
| Autor: | N. R. Burns, W. B. Gratzer |
|---|---|
| Rok vydání: | 1985 |
| Předmět: |
Erythrocytes
Calmodulin Ionophore chemistry.chemical_element Trifluoperazine Calcium Biochemistry Calcium in biology medicine Animals Humans Spectrin Binding site biology Chemistry Erythrocyte Membrane Microfilament Proteins Brain EPB41 Hydrogen-Ion Concentration Kinetics Microscopy Electron Scanning biology.protein Biophysics Cattle Carrier Proteins Protein Binding medicine.drug |
| Zdroj: | Biochemistry. 24:3070-3074 |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/bi00333a040 |
| Popis: | The binding of calmodulin to red cell membrane cytoskeletons and to purified spectrin from red cells and bovine brain spectrin (fodrin) has been examined. Under physiological solvent conditions binding can be measured by ultracentrifugal pelleting assays. The membrane cytoskeletons contained a single class of binding sites, with a concentration similar to that of spectrin dimers and an association constant of 1.5 X 10(5) M-1. Binding is calcium dependent and is suppressed by the calmodulin inhibitor trifluoperazine. The binding showed a marked dependence on ionic strength, with a maximum at 0.05 M, and a steep dependence on pH, with a maximum at pH 6.5. It was unaffected by 5 mM magnesium. An azidocalmodulin derivative, under the conditions of our experiments, did not label the spectrin-containing complex, although it could be used to demonstrate binding to fodrin. Binding of calmodulin to spectrin tetramers and fodrin in solution could be demonstrated by a pelleting assay after addition of F-actin. Calculations (which are necessarily rough) suggest that at the free calcium concentration prevailing in a normal red cell about 1 in 20 of the calmodulin binding sites in spectrin will be occupied; this proportion will rise rapidly with increasing intracellular calcium. To determine whether inhibition of calmodulin binding to red cell proteins disturbs the control of cell shape, as has been suggested, calcium ions were removed from the cell by addition of an ionophore and of ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid to the external medium. This did not affect the discoid shape. Trifluoperazine still induced stomatocytosis, exactly as in untreated cells.(ABSTRACT TRUNCATED AT 250 WORDS) |
| Databáze: | OpenAIRE |
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