Structure of the erythrocyte anion transport protein.
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Anion Exchange Protein 1, Erythrocyte↗
Biomedical subjects
R E Jenkins
Publications and source records attributed to R E Jenkins.
Structure of the anion-transport protein of the human erythrocyte membrane. Further studies on the fragments produced by proteolytic digestion.
The topology of the human erythrocyte membrane anion-transport protein (band 3) has been investigated by isolation and peptide 'mapping' of the major and minor fragments derived from proteolytic cleavage of the lactoperoxidase 125I-labelled protein in erythrocytes and erythrocyte membranes. The content, in each fragment, of lactoperoxidase 125I-labelled sites (which have a known location in the extracellular or cytoplasmic domain of the protein), together with the location of the sites of proteolytic cleavage yielding the fragments, has allowed us to determine the alignment of the fragments on the linear amino acid sequence and to infer the topology of the polypeptide in the membrane. The results suggest that a region in the C-terminal portion of the polypeptide forms part of the cytoplasmic domain of the protein in addition to a large N-terminal segment. The membrane-bound regions of the protein are located in the C-terminal two-thirds of the molecule. In this region the polypeptide chain traverses the membrane at least four times and an additional loop of polypeptide is either embedded in the membrane or also penetrates through it to the other surface. The location of the lectin receptors on the protein and the site of binding of an anion-transport inhibitor have also been studied.
Amino Acid Sequence↗
Ionic-strength-dependent changes in the structure of the major protein of the human erythrocyte membrane.
The effect of ionic strength on the proteolysis by trypsin of the major membrane-penetrating protein (polypeptide 3) in the erythrocyte membrane was studied. Both the intracellular and extracellular regions of the protein are susceptible to trypsin proteolysis under hypo-osmotic conditions, whereas under iso-osmotic conditions the extracellular region of the protein is resistant to trypsin, and the intracellular region yields only two cleavage products with trypsin. Studies of the fragments obtained from polypeptide 3 by trypsin digestion under iso-osmotic conditions of 'ghosts' radioiodinated with lactoperoxidase confirmed our earlier conclusions that the polypeptide chain of polypeptide 3 traverses the membrane twice. Ionic-strength-dependent changes were also observed in the incorporation of iodine by lactoperoxidase into the individual extracellular tyrosine sites of the protein. These results show that polypeptide 3 undergoes ionic-strength-dependent changes in structure.
Autoradiography↗
The structure of the major protein of the human erythrocyte membrane. Characterization of the intact protein and major fragments.
Polypeptide 3, the major membrane-penetrating protein of the human erythrocyte membrane, was characterized, together with two major fragments derived by specific proteolysis of the native protein in the membrane. One fragment (fragment 3f) was obtained from thermolysin cleavage in the extracellular region of the protein, and the other (fragment T1) was derived from tryptic cleavage in the intracellular region of the protein. The results of N- and C-terminal group analysis suggest that fragment 3f contains the N-terminal region of polypeptide 3 and fragment T1 contains the C-terminal part of the molecule. The carbohydrate contents of the polypeptides suggest that carbohydrates are present in three regions of the molecule, much of this carbohydrate being present in the C-terminal part of the molecule. This region of the protein also contains the receptors for concanavalin and the lectins from Phaseolus vulgaris and Ricinis communis, and our results suggest that there is heterogeneity in the carbohydrate chains present in the C-terminal region of polypeptide 3. These data are related to the folding of polypeptide 3 in the erythrocyte membrane.
Amino Acids↗
Abnormal carbohydrate composition of the major penetrating membrane protein of En(a-) human erythrocytes.
The major penetrating membrane glycoprotein (band 3) was isolated from En(a-) and normal human erythrocytes. The two proteins differed only in carbohydrate composition. Band 3 from En(a-) erythrocytes contained greater amounts of galactose and N-acetyl-glucosamine. The loss of the sialoglycoprotein sialotetrasaccharides in the En(a-) cell is not compensated by the appearance of these units in band 3 of En(a-) erythrocytes.
Acetylglucosamine↗
The major human erythrocyte membrane protein. Evidence for an S-shaped structure which traverses the membrane twice and contains a duplicated set of sites.
The structure of the major human erythrocyte membrane protein (protein E) was investigated by studying the products of proteolysis of the native protein in the membrane. The distribution and location of the tyrosine residues labelled by radioiodination by lactoperoxidase was determined. Proteolysis of the extracellular region of the protein by thermolysin released four tyrosine-containing peptides, all of which were also found to remain in the major fragment that is retained in the membrane. The presence of these duplicated sites in the extracellular region of the protein was confirmed by limited trypsin digestion of the intracellular region of the protein. Two groups of fragments were obtained. Both groups contained a set of the extracellular labelled sites, but they differed in containing distinct groups of intracellular sites, showing that the two sets of extracellular sites are linked by an intracellular region of the protein. The polypeptide chain thus traverses the membrane twice. An S-shaped model which is consistent with these data is proposed.
Cell Membrane↗
The organization of the major protein of the human erythrocyte membrane.
The enzyme lactoperoxidase was used to catalyse the radioiodination of membrane proteins in intact human erythrocytes and in erythrocyte ;ghosts'. Two major proteins of the erythrocyte membrane were isolated after iodination of these two preparations, and the peptide ;maps' of each protein so labelled were compared. Peptides from both proteins are labelled in the intact cell. In addition, further mobile peptides derived from one of the proteins are labelled only in the ;ghost' preparation. Various sealed ;ghost' preparations were also iodinated, lactoperoxidase being present only at either the cytoplasmic or extra-cellular surface of the membrane. The peptide ;maps' of protein E (the major membrane protein) labelled in each case were compared. Two discrete sets of labelled peptides were consistently found. One group is obtained when lactoperoxidase is present at the extra-cellular surface and the other group is found when the enzyme is accessible only to the cytoplasmic surface of the membrane. The results support the assumption that the organization of protein E in the membrane of the intact erythrocyte is unaltered on making erythrocyte ;ghosts'. They also confirm previous suggestions that both the sialoglycoprotein and protein E extend through the human erythrocyte membrane.
Autoradiography↗
Localized xanthomas in lymphedema praecox.
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Adolescent↗