Abstract
The extracellular surface of theα-chain ofTorpedo california acetylcholine receptor (AChR) was mapped for regions that are accessible to binding with antibodies against a panel of synthetic overlapping peptides which encompassed the entire extracellular parts of the chain. The binding of the antipeptide antibodies to membrane-bound AChR (mbAChR) and to isolated, soluble AChR. was determined. The specificity of each antiserum was narrowed down by determining the extent of its cross-reaction with the two adjacent peptides that overlap the immunizing peptide. With mbAChR, high antibody reactivity was obtained with antisera against peptidesα1–16,α89–104,α158–174,α262–276, andα388–408. Lower, but significant, levels of reactivity were obtained with antibodies against peptidesα67–82,α78–93,α100–115, andα111–126. On the other hand, free AChR bound high levels of antibodies against peptidesα34–49,α78–93,α134–150,α170–186, andα194–210. It also bound moderate levels of antibodies against peptidesα262–276 andα388–408. Low, yet significant, levels of binding were exhibited by antibodies against peptidesα45–60,α111–126, andα122–138. These binding studies, which enabled a comparison of the accessible regions in mbAChR and free AChR, revealed that the receptor undergoes considerable changes in conformation upon removal from the cell membrane. The exposed regions found here are discussed in relation to the functional sites of AChR (i.e., the acetylcholine binding site, the regions that are recognized by anti-AChR antibodies, T-cells and autoimmune responses and the regions that bind short and long neurotoxins).
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Abbreviations
- AChR:
-
acetylcholine receptor
- mbAChR:
-
membrane-bound AChR
- BSA:
-
bovine serum albumin
- BTX:
-
α-bungarotoxin
- EAMG:
-
experimental autoimmune myasthenia gravis
- MG:
-
myasthenia gravis
- PBS:
-
0.15 MNaCl in 0.01 M sodium phosphate buffer, pH 7.2.
References
Ashizawa, T., Ruan, K. H., Jinnai, K., and Atassi, M. Z. (1992).Mol. Immunol. 29, 1507–1514.
Atassi, M. Z. (1975).Immunochemistry 12, 423–438.
Atassi, M. Z. (1980).Mol. Cell. Biochem. 32, 21–44.
Atassi, M. Z. (1984).Eur. J. Biochem. 145, 1–20.
Atassi, M. Z. (1986).Methods in Enzym. (Langone, J. J., and van Vunakis, eds.), Vol. 121, Academic Press, New York, pp. 69–95.
Atassi, M. Z., and Habeeb, A. F. S. A. (1977). InImmunochemistry of Proteins (Atassi, M. Z., ed.), Vol. 2, Plenum Press, New York, pp. 177–264.
Atassi, M. Z., and Smith, J. A. (1978).Immunochemistry 15, 609–610.
Atassi, M. Z., and Young, C. R. (1985).Crit. Revs. Immunol. 5, 387–410.
Atassi, M. Z., Mulac-Jericevic, B., Yokoi, T., and Manshouri, T. (1987).Fed. Proc. 46, 2538–2547.
Atassi, M. Z., McDaniel, C. S., and Manshouri, T. (1988a).J. Prot. Chem. 7, 655–666.
Atassi, M. Z., Manshouri, T., and Yokoi, T. (1988b).FEBS Lett. 228, 295–300.
Atassi, M. Z., Ruan, K. H., Jinnai, K., Oshima, M., and Ashizawa, T. (1992).Proc. Natl. Acad. Sci. USA 89, 5852.
Boulter, J., Evans, K., Goldmann, D., Martin, G., Treco, D., Heinemann, D., and Patrick, J. (1986).Nature (London) 319, 368–374.
Changeux, J. P., Devillers-Thiery A., and Chemouilli, P. (1984).Science 225, 1335–1345.
Claudio, T., Ballivet, M., Patrick, J., and Heinemann, S. (1983).Proc. Natl. Acad. Sci. USA 80, 1111–1115.
Cohen, J. B., Sharp. S. D., and Liu, W. S. (1991).J. Biol. Chem. 266, 23354–23364.
David, C. S., and Atassi, M. Z. (1982).Adv. Exptl. Med. Biol. 150, 97–125.
Finer-Moore, J., and Stroud, R. M. (1984).Proc. Natl. Acad. Sci. USA 81, 155–159.
Froehner, S. C., and Rafto, M. A. (1979).Biochemistry 18, 301–307.
Guy, H. R. (1983).Biophys. 45, 249–261.
Haggerty, J. G., and Froehner, S. C. (1981),J. Biol. Chem. 256, 8294–8297.
Hopp, T. P., and Woods, K. R. (1981).Proc. Nat. Acad. Sci. USA 78, 3824–3828.
Karlin, A. (1980).Cell Surface and Neuronal Function (Colman, C. W., Poste, G. and Nicolson, G. L., eds.), Elsevier/North-Holland Biomedical Press, New York, pp. 191–260.
Kazim, A. L., and Atassi, M. Z. (1980).Biochem. J. 191, 261–264.
Kazim, A. L., and Atassi, M. Z. (1982).Biochem. J. 203, 201–208.
Lee, C. Y. (1979).Adv. Cytopharmacol. 3, 1–16.
Lennon, V. A., McCormick, D. J., Lambert, E. H., Griesmann, G. E., and Atassi, M. Z. (1985).Proc. Natl. Acad. Sci. (USA) 82, 8805–8809.
Maelicke, A., Fulpius, B. W., Klett, R. P., and Reich, E. (1977).J. Biol. Chem. 252, 4811–4830.
Meunier, J. C., Sealock, R., Olsen, R., and Changeux, J. P. (1974).Eur. J. Biochem. 45, 371–394.
McCormick, D. J., and Atassi, M. Z. (1984).Biochem. J. 224, 995–1000.
McDaniel, C. S., Manshouri, T., and Atassi, M. Z. (1987).J. Prot. Chem. 6, 455–461.
Moore, H. H., and Raftery, M. Z. (1979).Biochemistry 18, 1862–1867.
Mulac-Jericevic, B., and Atassi, M. Z. (1986).FEBS Lett. 199, 68–74.
Mulac-Jericevic, B., and Atassi, M. Z. (1987a).J. Prot. Chem. 6, 365–373.
Mulac-Jericevic, B., and Atassi, M. Z. (1987b).Biochem. J. 248, 847–852.
Mulac-Jericevic, B., Kurisaki, J., and Atassi, M. Z. (1987).Proc. Natl. Acad. Sci. USA 84, 3633–3637.
Mulac-Jericevic, B., Manshouri, T., Yokoi, T., and Atassi, M. Z. (1988).J. Prot. Chem. 7, 173–177.
Noda, M., Takahashi, H., Tanabe, T., Toyosato, M., Furutani, Y., Hirose, T., Asai, M., Inayama, S., Miyata, T., and Numa, S. (1982).Nature (London) 299, 793–797.
Noda, M., Takahashi, H., Tanabe, T., Toyosato, M., Kikyotani, S., Hirose, T., Asai, M., Takashima, H., Inayama, S., Miyata, T., and Numa, S. (1983a).Nature (London) 301, 251–255.
Noda, M., Takahashi, H., Tanabe, T., Toyosato, M., Kikyotani, Miyata, T., and Numa, S. (1983b).Nature (London) 302, 528–532.
Noda, M., Furutani, Y., Yakahashi, H., Toyosato, M., Tanabe, T., Schimizu, S., Kikyotani, S., Kayano, T., Hirose, T., Inoyama, S., and Numa, S. (1983c).Nature (London) 305, 818–823.
Okuda, K., Twining, S. S., David, C. S., and Atassi, M. Z. (1979).J. Immunol. 123, 182–188.
Oshima, M., and Atassi, M. Z. (1984).Immunol. Invest. 18, 841–851.
Oshima, M., Ashizawa, T., Pollack, M. A., and Atassi, M. Z. (1990).Eur. J. Immun. 20, 2563–2569.
Ruan, K.-H., Spurlino, J., Quiocho, F. A., and Atassi, M. Z. (1990).Proc. Natl. Acad. Sci. USA 87, 6156–6160.
Ruan, K.-H., Stiles, B. G., and Atassi, M. Z. (1991).Biochem. J. 274, 849–854.
Schmitz, H. E., Atassi, H., and Atassi, M. Z. (1983).Immunol. Commun. 12, 161–175.
Sobel, A., Weber, M., and Changeux, J. P. (1977).Eur. J. Biochem. 80, 215–224.
Tzartos, S. J., and Changeux, J. P. (1983).EMBO J. 2, 381–387.
Yokoi, T., Mulac-Jericevic, B., Kurisaki, J., and Atassi, M. Z. (1987).Eur. J. Immunol. 17, 1697–1702.
Young, C. R., and Atassi, M. Z. (1982).Immunol. Commun. 11, 9–16.
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Atassi, M.Z., Mulac-Jericevic, B. Mapping the extracellular topography of the α-chain in free and in membrane-bound acetylcholine receptor by antibodies against overlapping peptides spanning the entire extracellular parts of the chain. J Protein Chem 13, 37–47 (1994). https://doi.org/10.1007/BF01891991
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DOI: https://doi.org/10.1007/BF01891991