Selective alteration of erythrocyte deformabiliby by SH-reagents: evidence for an involvement of spectrin in membrane shear elasticity.
In order to elucidate the molecular basis of membrane shear elasticity, the effect of membrane protein modification by SH-reaents on the deformability of human erythrocytes was studied. Deformability was quuantified by measuring the elongation of erythrocytes subjected to viscometric flow in a transparent cone plate viscometer. Impermeable SH-reagents proved to have no mechanical effect. Many, but not all, permeable SH-reagents markedly decreased the elongation. Among these, bifunctional SH-reagents (e.g. diamide, tetrathionate and N, N' -p-phenylenedimaleimide) able to cross-link membrane SH-groups were more effective than monofunctional SH-reagents (e.g. N-ethylmaleimide and ethacrynic acid). The bifunctional SH-reagents produced a 50% decrease of elongation after modification of less than 5% of the membrane SH-groups. In contrast, for a comparable effect, more than 20% of the SH-groups had to be modified by the monofunctional reagents. The effect of SH-oxidizing agents was fully reversible after treatment with disulfide-reducing agents. All bifunctional SH-reagents induced a dimerization of a small fraction of spectrin. Anaalysis of the distribution of the diamide-induced disulfide bonds among the various membrane protein fractions showed that this agent preferentially acts on the spectrin polypeptides. The results provide direct experimental evidence that the native arrangement of spectrin is essential for the shear resistance of the erythrocyte membrane and that introduction of small numbers of intermolecular cross-links as well as modification within the molecule lead to a rapid loss of this function.
D003958:Diamide / Q000494:pharmacology; D004220:Disulfides / Q000378:metabolism; D004548:Elasticity; D004910:Erythrocyte Membrane / Q000187:drug effects* / Q000502:physiology / Q000648:ultrastructure; D004912:Erythrocytes / Q000187:drug effects*; D006801:Humans; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000502:physiology*; D013049:Spectrin / Q000502:physiology*; D013439:Sulfhydryl Reagents / Q000494:pharmacology*
D004220:Disulfides; D008565:Membrane Proteins; D013439:Sulfhydryl Reagents; D003958:Diamide; D013049:Spectrin