Identification of the Membrane Attachment Sites for Protein 4.1 in the Human Erythrocyte (∗)

The nature of the membrane attachment site(s) for protein 4.1 in the human erythrocyte membrane has yet to be fully elucidated. In this paper we show that the major attachment site is glycophorin (GP) C/D, and that purified protein 4.1 can bind to two distinct sites on glycophorin C/D. One of these...

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Bibliographic Details
Published in:The Journal of biological chemistry 1995-03, Vol.270 (10), p.5360-5366
Main Authors: Hemming, Nicola J., Anstee, David J., Staricoff, Marcelo A., Tanner, Michael J.A., Mohandas, Narla
Format: Article
Language:English
Subjects:
Binding Sites
Chymotrypsin
Cytoskeletal Proteins
Electrophoresis, Polyacrylamide Gel
Enzyme-Linked Immunosorbent Assay
Erythrocyte Membrane - metabolism
Glycophorins - chemistry
Glycophorins - isolation & purification
Glycophorins - metabolism
Humans
Kinetics
Membrane Proteins - chemistry
Membrane Proteins - isolation & purification
Membrane Proteins - metabolism
Models, Structural
Neuropeptides
Protein Conformation
Trypsin
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Summary:The nature of the membrane attachment site(s) for protein 4.1 in the human erythrocyte membrane has yet to be fully elucidated. In this paper we show that the major attachment site is glycophorin (GP) C/D, and that purified protein 4.1 can bind to two distinct sites on glycophorin C/D. One of these interactions is direct, involving residues 82-98 on glycophorin C (61-77 on glycophorin D), while the other interaction is mediated by p55. We have localized the binding site for p55 on glycophorin C to residues 112-128 (glycophorin D 91-107). We also provide evidence that band 3 is an additional, minor, protein 4.1 binding site. The binding sites for band 3, glycophorin C/D, and p55 are all located within the 30-kDa domain of protein 4.1. We estimate that the relative utilization of the three sites in normal membranes comprises 40% to p55, 40% to GPC/D, and 20% to band 3. The same region of protein 4.1 binds GPC/D and band 3, while the p55 binding site is distinct. The interactions involving protein 4.1 with p55 and p55 with GPC/D are of high affinity (nM), while those involving GPC/D and band 3 are 100-fold lower (μM). These results suggest that the most significant interactions between protein 4.1 and the membrane are those involving p55.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.270.10.5360