Ankyrin is an essential link between cytoskeletal proteins, such as spectrin, and membrane bound proteins, such as protein 3, the erythrocyte anion exchanger. Although the amino acid structure of human ankyrin is known, the functional regions have been only partially defined. Sequence comparisons between mouse and human ankyrin offer one mechanism of identifying highly conserved regions that probably have functional significance. We report the isolation and sequencing of a series of overlapping murine erythroid ankyrin (Ank-1) cDNAs from spleen and reticulocyte libraries (total span 6238 bp) and identify potentially important regions of murine-human reticulocyte ankyrin homology. Comparison of the predicted peptide sequences of mouse and human erythroid ankyrins shows that these ankyrins are highly conserved in both the N-terminal, protein 3 binding domain (96% amino acid identity) and in the central spectrin-binding domain (97% identity), but differ in the C-terminal regulatory domain (79% identity). However, the C-terminal regulatory domain contains two regions of peptide sequence that are perfectly conserved. We postulate these regions are important in the regulatory functions of this domain.
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Bennett, V.: Purification of an active proteolytic fragment of the membrane attachment site for human erythrocyte spectrin. J Biol Chem 253: 2293–2299, 1978.
Bennett, V.: The membrane skeleton of human erythrocytes and its implications for more complex cells. Annu Rev Biochem 54: 273–304, 1985.
Bennett, V.: Spectrin-based membrane skeleton: a multipotential adaptor between plasma membrane and cytoplasm. Physiol Rev 70: 1029–1065, 1990.
Bennett, V. and Davis, J.: Erythrocyte ankyrin: immunoreactive analogues are associated with mitotic structures in cultured cells and with microtubules from brain. Proc Natl Acad Sci USA 78: 7550–7554, 1981.
Bennett, V. and Stenbuck, P.J.: The membrane attachment protein for spectrin is associated with band 3 in human erythrocyte membranes. Nature 280: 468–473, 1979.
Bennett, V. and Stenbuck, P.J.: Association between ankyrin and the cytoplasmic domain of band 3 isolated from human erythrocyte membrane. J Biol Chem 255: 6424–6432, 1980.
Birkenmeier, C.S., Blood, M.L., and Barker, J.E.: Preparation of a reticulocyte cDNA library from mouse: isolation of multiple overlapping clones for alpha- and beta-spectrin and ankyrin by cDNA walking. Blood 76 [Suppl 1]: 4a, 1990 (abstr).
Bodine, D.B., Birkenmeier, C.S., and Barker, J.E.: Spectrin deficient inherited hemolytic anemias in the mouse: characterization by spectrin synthesis and mRNA activity in reticulocytes. Cell 37 721–729, 1984.
Chomczynski, P. and Sacchi, N.: Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162: 156–159, 1987.
Chui, D.H.K., Patterson, M., and Bayly, S.T.: Unequal alpha and beta globin mRNA in reticulocytes of normal and mutant f/f fetal mice. Br J Haematol 44: 431–439, 1980.
Costa, F.F., Agre, P., Watkins, P.C., Winkelmann, J.C., Tang, T.K., John, K.M., Lux, S.E., and Forget, B.G.: Linkage of dominant hereditary spherocytosis to the gene for the erythrocyte membrane-skeleton protein ankyrin. N Engl J Med 323: 1046–1050, 1990.
Davis, J.Q. and Bennett, V.: Brain ankyrin. A membrane-associated protein with binding sites for spectrin, tubulin, and the cytoplasmic domain of the erythrocyte anion channel. J Biol Chem 259: 13550–13559, 1984.
Davis, L.H. and Bennett, V.: Mapping the binding sites for human erythrocyte ankyrin for the anion exchanger and spectrin. J Biol Chem 265: 10589–19596, 1990.
Davis, J., Davis, L., and Bennett, V.: Diversity in membrane binding sites of ankyrins. Brain ankyrin, erythrocyte ankyrin, and processed erythrocyte ankyrin associate with distinct sites in kidney melanosomes. J Biol Chem 264: 6417–6426, 1989.
Bel Sal, G., Manfioletti, G., and Schneider, C.: The CTAB-DNA precipitation method: a common mini-scale preparation of template DNA from phagemids, phages or plasmids suitable for sequencing. Biotechniques 7: 514–520, 1989.
Devereux, J., Haeberli, P., and Smithies, O.: A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res 12: 387–395, 1984.
Feinberg, A.P. and Vogelstein, B.: A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 137: 266, 1984.
Gray, P.W., Barrett, K., Chantry, D., Turner, M., and Feldman, M.: Cloning of tumor necrosis factor (TNF) receptor cDNA and expression of recombinant soluble TNF-binding protein. Proc Natl Acad Sci USA 87: 7380–7384, 1990.
Hall, T.G. and Bennett, V.: Regulatory domains of erythrocyte ankyrin. J Biol Chem 262: 10537–10545, 1987.
Hanspal, M., Yoon, S., Yu, H., Hanspal, J.S., Lambert, S., Palek, J., and Prchal, J.T.: Molecular basis of spectrin and ankyrin deficiencies in severe hereditary spherocytosis: evidence implicating a primary defect of ankyrin. Blood 77: 165–173, 1991.
Hargreaves, W.R., Giedd, K.N., Verkleij, A., and Branton, D.: Reassociation of ankyrin with band 3 in erythrocyte membranes and in lipid vesicles. J Biol Chem 255: 11965–11972, 1980.
Henikoff, S.: Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing. Gene 28: 351–359, 1984.
Howard, S.T., Chan, Y.S., and Smith, G.L.: Vaccinia virus homologues of the Shope fibroma virus inverted terminal repeat proteins and a continuous ORF related to the tumor necrosis factor receptor family. Virology 180: 633–647, 1991.
Kozak, M.: Comparison of initiation of protein synthesis in procaryotes, eucaryotes and organelles. Microbiol Rev 47: 1–45, 1983.
Kozak, M.: Compilation and analysis of sequences upstream from the translation start site in eucaryotic mRNAs. Nucleic Acids Res 12: 857–914, 1984.
Kozak, M.: An analysis of 5′-noncoding sequences from 699 vertebrate messenger RNAs. Nucleic Acids Res 15: 8125–8148, 1987.
Lambert, S., Yu, H., Prchal, J.T., Lawler, J., Ruff, P., Speicher, D., Cheung, M.C., Kan, Y.W., and Palek, J.: cDNA sequence for human erythrocyte ankyrin. Proc Natl Acad Sci USA 87: 1730–1734, 1990.
Low, P.S., Williardson, B.M., Mohandas, N., Rossi, M., and Shohet, S.: Contribution of the band 3-ankyrin interaction to membrane mechanical stability. Blood 77: 1581–1586, 1991.
Lux, S.E., John, K.M., and Bennett, V.: Analysis of cDNA for human erythrocyte ankyrin indicates a repeated structure with homology to tissue-differentiation and cell-cycle proteins. Nature 344: 36–42, 1990a.
Lux, S.E., Tse, W.T., Menninger, J.C., John, K.M., Harris, P., Shalev, O., Chilcote, R.R., Marchesi, S.L., Watkins, P.C., Bennett, V., McIntosh, S., Collins, F.C., Francke, U., Ward, D.C., and Forget, B.G.: Hereditary spherocytosis associated with deletion of human erythrocyte ankyrin gene on chromosome 8. Nature 345: 736–739, 1990b.
Otto, E., Kunimoto, M., McLaughlin, T., and bennett, V.: Isolation and characterization of cDNAs encoding human brain ankyrins reveal a family of alternatively spliced genes. J Cell Biol 114: 241–253, 1991.
Peters, L.L., Birkenmeier, C.S., Bronson, R.T., White, R.A., Lux, S.E., Otto, E., Bennett, V., and Barker, J.E.: Purkinje cell degeneration associated with erythroid ankyrin deficiency in nb/nb mice. J Cell Biol 114: 114–121, 1991.
Platt, O.S., Falcone, J.F., and Lux, S.E.: A 12 kD tryptic peptide of ankyrin retains spectrin binding capacity. Blood 74 [Suppl 1]: 104a, 1989 (abstr).
Sambrook, J., Fritsch, T. and Maniatis, T.: Molecular Cloning: a Laboratory Manual, 2nd edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989.
Sanger, F., Nicklen, S., and Coulson, A.R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74: 5463–5467, 1977.
Short, J.M., Fernandez, J.M., Sorge, J.A., and Huse, W.D.: Lambda ZAP: a bacteriophage lambda expression vector with in vivo excision properties. Nucleic Acids Res 16: 7583–7600, 1988.
Toneguzzo, F., Glynn, S., Levi, E., Mjolsness, S., and Hayday, A.: Use of a chemically modified T7 DNA polymerase for manual and automated sequencing of supercoiled DNA. Biotechniques 6: 460–469, 1988.
Wallin, R., Culp, E.N., Coleman, D.B., and Goodman, S.R.: A structural model of human erythrocyte band 2.1: alignment of chemical and functional domains. Proc Natl Acad Sci USA 81: 4095–4099, 1984.
Weaver, D.C. and Marchesi, V.T.: The structural basis of ankyrin function. I. Identification of two structural domains. J Biol Chem 259: 6165–6169, 1984a.
Weaver, D.C. and Marchesi, V.T.: The structural basis of ankyrin function. II. Identification of two functional domains. J Biol Chem 259: 6170–6175, 1984b.
White, R.A., Birkenmeier, C.S., Lux, S.E., and Barker, J.E.: Ankyrin and the hemolytic anemia mutation, nb, map to mouse chromosome 8: presence of the nb allele is associated with a truncated erythrocyte ankyrin. Proc Natl Acad Sci USA 87: 3117–3121, 1990.
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White, R.A., Birkenmeier, C.S., Peters, L.L. et al. Murine erythrocyte ankyrin cDNA: highly conserved regions of the regulatory domain. Mammalian Genome 3, 281–285 (1992). https://doi.org/10.1007/BF00292156
- Binding Domain
- Anion Exchanger
- Peptide Sequence
- Amino Acid Identity
- Acid Identity