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. 1991 Oct 1;279(Pt 1):203–212. doi: 10.1042/bj2790203

Definition of surface-exposed epitopes on the (Ca(2+)-Mg2+)-ATPase of sarcoplasmic reticulum.

R E Tunwell 1, J W Conlan 1, I Matthews 1, J M East 1, A G Lee 1
PMCID: PMC1151567  PMID: 1718260

Abstract

Epitopes for monoclonal antibodies binding to the native (Ca(2+)-Mg2+)-ATPase have been defined by studying binding to sets of hexameric peptides synthesized on plastic pegs. Epitopes have been confirmed by demonstrating the binding of anti-peptide antibodies to the ATPase. A method is presented for definition of surface-exposed epitopes using polyclonal antibodies. Three surface-exposed epitopes have been defined in the nucleotide-binding domain of the ATPase, suggesting considerable surface exposure of this region. Other surface-exposed epitopes have been located in the region of the fourth stalk domain.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Atassi M. Z. Antigenic structure of myoglobin: the complete immunochemical anatomy of a protein and conclusions relating to antigenic structures of proteins. Immunochemistry. 1975 May;12(5):423–438. doi: 10.1016/0019-2791(75)90010-5. [DOI] [PubMed] [Google Scholar]
  2. Bishop J. E., Squier T. C., Bigelow D. J., Inesi G. (Iodoacetamido)fluorescein labels a pair of proximal cysteines on the Ca2+-ATPase of sarcoplasmic reticulum. Biochemistry. 1988 Jul 12;27(14):5233–5240. doi: 10.1021/bi00414a043. [DOI] [PubMed] [Google Scholar]
  3. Brandl C. J., Green N. M., Korczak B., MacLennan D. H. Two Ca2+ ATPase genes: homologies and mechanistic implications of deduced amino acid sequences. Cell. 1986 Feb 28;44(4):597–607. doi: 10.1016/0092-8674(86)90269-2. [DOI] [PubMed] [Google Scholar]
  4. Clarke D. M., Loo T. W., Inesi G., MacLennan D. H. Location of high affinity Ca2+-binding sites within the predicted transmembrane domain of the sarcoplasmic reticulum Ca2+-ATPase. Nature. 1989 Jun 8;339(6224):476–478. doi: 10.1038/339476a0. [DOI] [PubMed] [Google Scholar]
  5. Colyer J., Mata A. M., Lee A. G., East J. M. Effects on ATPase activity of monoclonal antibodies raised against (Ca2+ + Mg2+)-ATPase from rabbit skeletal muscle sarcoplasmic reticulum and their correlation with epitope location. Biochem J. 1989 Sep 1;262(2):439–447. doi: 10.1042/bj2620439. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Fasman G. D., Gilbert W. A. The prediction of transmembrane protein sequences and their conformation: an evaluation. Trends Biochem Sci. 1990 Mar;15(3):89–92. doi: 10.1016/0968-0004(90)90187-g. [DOI] [PubMed] [Google Scholar]
  7. Fields G. B., Noble R. L. Solid phase peptide synthesis utilizing 9-fluorenylmethoxycarbonyl amino acids. Int J Pept Protein Res. 1990 Mar;35(3):161–214. doi: 10.1111/j.1399-3011.1990.tb00939.x. [DOI] [PubMed] [Google Scholar]
  8. Froud R. J., East J. M., Rooney E. K., Lee A. G. Binding of long-chain alkyl derivatives to lipid bilayers and to (Ca2+-Mg2+)-ATPase. Biochemistry. 1986 Nov 18;25(23):7535–7544. doi: 10.1021/bi00371a042. [DOI] [PubMed] [Google Scholar]
  9. Geysen H. M., Rodda S. J., Mason T. J., Tribbick G., Schoofs P. G. Strategies for epitope analysis using peptide synthesis. J Immunol Methods. 1987 Sep 24;102(2):259–274. doi: 10.1016/0022-1759(87)90085-8. [DOI] [PubMed] [Google Scholar]
  10. Gould G. W., Colyer J., East J. M., Lee A. G. Silver ions trigger Ca2+ release by interaction with the (Ca2+-Mg2+)-ATPase in reconstituted systems. J Biol Chem. 1987 Jun 5;262(16):7676–7679. [PubMed] [Google Scholar]
  11. Green N. M. ATP-driven cation pumps: alignment of sequences. Biochem Soc Trans. 1989 Dec;17(6):972–972. doi: 10.1042/bst0170972. [DOI] [PubMed] [Google Scholar]
  12. Green N., Alexander H., Olson A., Alexander S., Shinnick T. M., Sutcliffe J. G., Lerner R. A. Immunogenic structure of the influenza virus hemagglutinin. Cell. 1982 Mar;28(3):477–487. doi: 10.1016/0092-8674(82)90202-1. [DOI] [PubMed] [Google Scholar]
  13. Gutierrez-Merino C., Munkonge F., Mata A. M., East J. M., Levinson B. L., Napier R. M., Lee A. G. The position of the ATP binding site on the (Ca2+ + Mg2+)-ATPase. Biochim Biophys Acta. 1987 Feb 26;897(2):207–216. doi: 10.1016/0005-2736(87)90417-2. [DOI] [PubMed] [Google Scholar]
  14. Hall C. N., Jamison M. H., MacLennan I. Complications of non-absorbable ligatures in thyroid surgery. Br J Clin Pract. 1985 Aug;39(8):307–309. [PubMed] [Google Scholar]
  15. Jähnig F. Structure predictions of membrane proteins are not that bad. Trends Biochem Sci. 1990 Mar;15(3):93–95. doi: 10.1016/0968-0004(90)90188-h. [DOI] [PubMed] [Google Scholar]
  16. Jørgensen P. L., Andersen J. P. Structural basis for E1-E2 conformational transitions in Na,K-pump and Ca-pump proteins. J Membr Biol. 1988 Jul;103(2):95–120. doi: 10.1007/BF01870942. [DOI] [PubMed] [Google Scholar]
  17. Kawakita M., Yamashita T. Reactive sulfhydryl groups of sarcoplasmic reticulum ATPase. III. Identification of cysteine residues whose modification with N-ethylmaleimide leads to loss of the Ca2+-transporting activity. J Biochem. 1987 Jul;102(1):103–109. doi: 10.1093/oxfordjournals.jbchem.a122021. [DOI] [PubMed] [Google Scholar]
  18. Kison R., Meyer H. E., Schoner W. Characterization of a cysteine-containing peptide after affinity labelling of Ca2+-ATPase of sarcoplasmic reticulum with the disulfide of 3'(2')-O-biotinyl-thioinosine triphosphate. Eur J Biochem. 1989 May 1;181(2):503–511. doi: 10.1111/j.1432-1033.1989.tb14752.x. [DOI] [PubMed] [Google Scholar]
  19. Laver W. G., Air G. M., Webster R. G., Smith-Gill S. J. Epitopes on protein antigens: misconceptions and realities. Cell. 1990 May 18;61(4):553–556. doi: 10.1016/0092-8674(90)90464-p. [DOI] [PubMed] [Google Scholar]
  20. MacLennan D. H., Brandl C. J., Korczak B., Green N. M. Amino-acid sequence of a Ca2+ + Mg2+-dependent ATPase from rabbit muscle sarcoplasmic reticulum, deduced from its complementary DNA sequence. Nature. 1985 Aug 22;316(6030):696–700. doi: 10.1038/316696a0. [DOI] [PubMed] [Google Scholar]
  21. Maruyama K., MacLennan D. H. Mutation of aspartic acid-351, lysine-352, and lysine-515 alters the Ca2+ transport activity of the Ca2+-ATPase expressed in COS-1 cells. Proc Natl Acad Sci U S A. 1988 May;85(10):3314–3318. doi: 10.1073/pnas.85.10.3314. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Matthews I., Colyer J., Mata A. M., Green N. M., Sharma R. P., Lee A. G., East J. M. Evidence for the cytoplasmic location of the N- and C-terminal segments of sarcoplasmic reticulum (Ca2+-Mg2+)-ATPase. Biochem Biophys Res Commun. 1989 Jun 15;161(2):683–688. doi: 10.1016/0006-291x(89)92653-3. [DOI] [PubMed] [Google Scholar]
  23. Merrifield B. Solid phase synthesis. Science. 1986 Apr 18;232(4748):341–347. doi: 10.1126/science.3961484. [DOI] [PubMed] [Google Scholar]
  24. Mitchinson C., Wilderspin A. F., Trinnaman B. J., Green N. M. Identification of a labelled peptide after stoicheiometric reaction of fluorescein isothiocyanate with the Ca2+ -dependent adenosine triphosphatase of sarcoplasmic reticulum. FEBS Lett. 1982 Sep 6;146(1):87–92. doi: 10.1016/0014-5793(82)80710-2. [DOI] [PubMed] [Google Scholar]
  25. Molnar E., Seidler N. W., Jona I., Martonosi A. N. The binding of monoclonal and polyclonal antibodies to the Ca2(+)-ATPase of sarcoplasmic reticulum: effects on interactions between ATPase molecules. Biochim Biophys Acta. 1990 Apr 13;1023(2):147–167. doi: 10.1016/0005-2736(90)90410-p. [DOI] [PubMed] [Google Scholar]
  26. Munkonge F., East J. M., Lee A. G. Positions of the sites labeled by N-cyclohexyl-N'-(4-dimethylamino-1-naphthyl)carbodiimide on the (Ca2+ + Mg2+)-ATPase. Biochim Biophys Acta. 1989 Feb 13;979(1):113–120. doi: 10.1016/0005-2736(89)90530-0. [DOI] [PubMed] [Google Scholar]
  27. Novotny J., Bruccoleri R. E., Saul F. A. On the attribution of binding energy in antigen-antibody complexes McPC 603, D1.3, and HyHEL-5. Biochemistry. 1989 May 30;28(11):4735–4749. doi: 10.1021/bi00437a034. [DOI] [PubMed] [Google Scholar]
  28. Obara M., Suzuki H., Kanazawa T. Conformational changes in the vicinity of the N-iodoacetyl-N'-(5-sulfo-1-naphthyl)ethylenediamine attached to the specific thiol of sarcoplasmic reticulum Ca2+-ATPase throughout the catalytic cycle. J Biol Chem. 1988 Mar 15;263(8):3690–3697. [PubMed] [Google Scholar]
  29. Ohta T., Nagano K., Yoshida M. The active site structure of Na+/K+-transporting ATPase: location of the 5'-(p-fluorosulfonyl)benzoyladenosine binding site and soluble peptides released by trypsin. Proc Natl Acad Sci U S A. 1986 Apr;83(7):2071–2075. doi: 10.1073/pnas.83.7.2071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Ovchinnikov YuA, Arzamazova N. M., Arystarkhova E. A., Gevondyan N. M., Aldanova N. A., Modyanov N. N. Detailed structural analysis of exposed domains of membrane-bound Na+,K+-ATPase. A model of transmembrane arrangement. FEBS Lett. 1987 Jun 15;217(2):269–274. doi: 10.1016/0014-5793(87)80676-2. [DOI] [PubMed] [Google Scholar]
  31. Ovchinnikov YuA, Luneva N. M., Arystarkhova E. A., Gevondyan N. M., Arzamazova N. M., Kozhich A. T., Nesmeyanov V. A., Modyanov N. N. Topology of Na+,K+-ATPase. Identification of the extra- and intracellular hydrophilic loops of the catalytic subunit by specific antibodies. FEBS Lett. 1988 Jan 25;227(2):230–234. doi: 10.1016/0014-5793(88)80904-9. [DOI] [PubMed] [Google Scholar]
  32. Petithory J. R., Jencks W. P. Phosphorylation of the calcium adenosinetriphosphatase of sarcoplasmic reticulum: rate-limiting conformational change followed by rapid phosphoryl transfer. Biochemistry. 1986 Aug 12;25(16):4493–4497. doi: 10.1021/bi00364a006. [DOI] [PubMed] [Google Scholar]
  33. Pick U. Interaction of fluorescein isothiocyanate with nucleotide-binding sites of the Ca-ATPase from sarcoplasmic reticulum. Eur J Biochem. 1981 Dec;121(1):187–195. doi: 10.1111/j.1432-1033.1981.tb06448.x. [DOI] [PubMed] [Google Scholar]
  34. Reithmeier R. A., MacLennan D. H. The NH2 terminus of the (Ca2+ + Mg2+)-adenosine triphosphatase is located on the cytoplasmic surface of the sarcoplasmic reticulum membrane. J Biol Chem. 1981 Jun 25;256(12):5957–5960. [PubMed] [Google Scholar]
  35. Scott T. L. Distances between the functional sites of the (Ca2+ + Mg2+)-ATPase of sarcoplasmic reticulum. J Biol Chem. 1985 Nov 25;260(27):14421–14423. [PubMed] [Google Scholar]
  36. Serrano R., Portillo F. Catalytic and regulatory sites of yeast plasma membrane H(+)-ATPase studied by directed mutagenesis. Biochim Biophys Acta. 1990 Jul 25;1018(2-3):195–199. doi: 10.1016/0005-2728(90)90247-2. [DOI] [PubMed] [Google Scholar]
  37. Serrano R. Structure and function of proton translocating ATPase in plasma membranes of plants and fungi. Biochim Biophys Acta. 1988 Feb 24;947(1):1–28. doi: 10.1016/0304-4157(88)90017-2. [DOI] [PubMed] [Google Scholar]
  38. Shull G. E., Schwartz A., Lingrel J. B. Amino-acid sequence of the catalytic subunit of the (Na+ + K+)ATPase deduced from a complementary DNA. Nature. 1985 Aug 22;316(6030):691–695. doi: 10.1038/316691a0. [DOI] [PubMed] [Google Scholar]
  39. Squier T. C., Bigelow D. J., Garcia de Ancos J., Inesi G. Localization of site-specific probes on the Ca-ATPase of sarcoplasmic reticulum using fluorescence energy transfer. J Biol Chem. 1987 Apr 5;262(10):4748–4754. [PubMed] [Google Scholar]
  40. Stokes D. L., Green N. M. Structure of CaATPase: electron microscopy of frozen-hydrated crystals at 6 A resolution in projection. J Mol Biol. 1990 Jun 5;213(3):529–538. doi: 10.1016/s0022-2836(05)80213-x. [DOI] [PubMed] [Google Scholar]
  41. Taylor W. R., Green N. M. The predicted secondary structures of the nucleotide-binding sites of six cation-transporting ATPases lead to a probable tertiary fold. Eur J Biochem. 1989 Jan 15;179(1):241–248. doi: 10.1111/j.1432-1033.1989.tb14547.x. [DOI] [PubMed] [Google Scholar]
  42. Tramontano A., Chothia C., Lesk A. M. Structural determinants of the conformations of medium-sized loops in proteins. Proteins. 1989;6(4):382–394. doi: 10.1002/prot.340060405. [DOI] [PubMed] [Google Scholar]
  43. Tunwell R. E., O'Connor C. D., Mata A. M., East J. M., Lee A. G. Mapping epitopes on the (Ca(2+)-Mg2+)-ATPase of sarcoplasmic reticulum using fusion proteins. Biochim Biophys Acta. 1991 Apr 9;1073(3):585–592. doi: 10.1016/0304-4165(91)90234-8. [DOI] [PubMed] [Google Scholar]
  44. Van Regenmortel M. H. The concept and operational definition of protein epitopes. Philos Trans R Soc Lond B Biol Sci. 1989 Jun 12;323(1217):451–466. doi: 10.1098/rstb.1989.0023. [DOI] [PubMed] [Google Scholar]
  45. Xu K. Y., Kyte J. Nucleophilic behavior of lysine-501 of the alpha-polypeptide of sodium and potassium ion activated adenosinetriphosphatase consistent with a role in binding adenosine triphosphate. Biochemistry. 1989 Apr 4;28(7):3009–3017. doi: 10.1021/bi00433a041. [DOI] [PubMed] [Google Scholar]
  46. Yamamoto H., Tagaya M., Fukui T., Kawakita M. Affinity labeling of the ATP-binding site of Ca2+-transporting ATPase of sarcoplasmic reticulum by adenosine triphosphopyridoxal: identification of the reactive lysyl residue. J Biochem. 1988 Mar;103(3):452–457. doi: 10.1093/oxfordjournals.jbchem.a122291. [DOI] [PubMed] [Google Scholar]

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