The redox state of cysteines 201 and 317 of the erythrocyte anion exchanger is critical for ankyrin binding

B. J.M. Thevenin, B. M. Willardson, P. S. Low

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58 Scopus citations


Previous studies have demonstrated that modification of erythrocyte membrane cysteine residues via disulfide cross-bridging or direct derivatization with thiol reagents promotes massive morphological, rheological, and structural changes in the cell. To determine whether disruption of the band 3-ankyrin interaction, the major membrane-cytoskeletal linkage, might contribute to the above lesions, we quantitatively measured the band 3-ankyrin interaction following modification of Cys-201 and/or Cys-317 of the cytoplasmic domain of band 3. It was observed that irreversible alkylating agents (e.g. N-ethylmaleimide or iodoacetamide and its derivatives), reversible derivatizing compounds (.eg.g. p-chloromercuribenzenesulfonate or glutathione), and native disulfide bond formation all blocked the ankyrin interaction. Comparison of the extent of sulfhydryl modification with the degree of inhibition of ankyrin binding further confirmed that cysteine modification was directly responsible for the inhibition. However, analysis of the site of sulfhydryl derivatization revealed that inhibition of ankyrin binding could be initiated in some cases with derivatization of Cys-201, while in other cases obstruction of Cys-317 appeared to be essential. This apparent discrepancy was resolved by demonstrating that Cys-201 of one strand of the cytoplasmic domain of band 3 dimer could disulfide bond with Cys-317 of the opposite strand, thus demonstrating that all four cysteines of the band 3 dimer are clustered at the interface between subunits. We argue that derivatization or disulfide cross-linking of these cysteines can block ankyrin binding by both conformational and steric mechanisms.

Original languageEnglish (US)
Pages (from-to)15886-15892
Number of pages7
JournalJournal of Biological Chemistry
Issue number27
StatePublished - 1989

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


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