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. 1983 Mar;80(6):1506–1510. doi: 10.1073/pnas.80.6.1506

Peptide antibody specific for the amino terminus of skeletal muscle alpha-actin.

J C Bulinski, S Kumar, K Titani, S D Hauschka
PMCID: PMC393630  PMID: 6572911

Abstract

The NH2-terminal peptide of skeletal muscle alpha-actin (S alpha N peptide), which contains a primary sequence unique to this actin isozyme, was used to prepare an isozyme-specific peptide antibody. S alpha N peptide was purified from chicken breast muscle actin by preparative reverse-phase HPLC and was coupled to hemocyanin. This complex was used to immunize rabbits in order to elicit actin antibodies specific for the skeletal muscle alpha-actin isozyme. The antibody obtained, called S alpha N antibody, was reactive with S alpha N peptide and with skeletal muscle alpha-actin as well as with cardiac muscle alpha-actin. S alpha N antibody did not react with either of the actin isozymes present in smooth muscle (smooth muscle alpha and gamma) or in brain (nonmuscle beta and gamma). S alpha N antibody was used to detect muscle-specific actin in differentiating mouse and human myoblasts by using immunoblots of myoblast extracts and immunofluorescent staining of fixed cells.

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Selected References

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

  1. Arnon R., Sela M. Antibodies to a unique region in lysozyme provoked by a synthetic antigen conjugate. Proc Natl Acad Sci U S A. 1969 Jan;62(1):163–170. doi: 10.1073/pnas.62.1.163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Atherton B. T., Hynes R. O. A difference between plasma and cellular fibronectins located with monoclonal antibodies. Cell. 1981 Jul;25(1):133–141. doi: 10.1016/0092-8674(81)90237-3. [DOI] [PubMed] [Google Scholar]
  3. Audibert F., Jolivet M., Chedid L., Alouf J. E., Boquet P., Rivaille P., Siffert O. Active antitoxic immunization by a diphtheria toxin synthetic oligopeptide. Nature. 1981 Feb 12;289(5798):593–594. doi: 10.1038/289593a0. [DOI] [PubMed] [Google Scholar]
  4. Bulinski J. C., Borisy G. G. Immunofluorescence localization of HeLa cell microtubule-associated proteins on microtubules in vitro and in vivo. J Cell Biol. 1980 Dec;87(3 Pt 1):792–801. doi: 10.1083/jcb.87.3.792. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Erlanger B. F. The preparation of antigenic hapten-carrier conjugates: a survey. Methods Enzymol. 1980;70(A):85–104. doi: 10.1016/s0076-6879(80)70043-5. [DOI] [PubMed] [Google Scholar]
  6. Garrels J. I., Gibson W. Identification and characterization of multiple forms of actin. Cell. 1976 Dec;9(4 Pt 2):793–805. doi: 10.1016/0092-8674(76)90142-2. [DOI] [PubMed] [Google Scholar]
  7. Gordon D. J., Boyer J. L., Korn E. D. Comparative biochemistry of non-muscle actins. J Biol Chem. 1977 Nov 25;252(22):8300–8309. [PubMed] [Google Scholar]
  8. Gordon D. J., Eisenberg E., Korn E. D. Characterization of cytoplasmic actin isolated from Acanthamoeba castellanii by a new method. J Biol Chem. 1976 Aug 10;251(15):4778–4786. [PubMed] [Google Scholar]
  9. Hauschka S. D. Clonal analysis of vertebrate myogenesis. II. Environmental influences upon human muscle differentiation. Dev Biol. 1974 Apr;37(2):329–344. doi: 10.1016/0012-1606(74)90153-5. [DOI] [PubMed] [Google Scholar]
  10. Hopp T. P., Woods K. R. Prediction of protein antigenic determinants from amino acid sequences. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3824–3828. doi: 10.1073/pnas.78.6.3824. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hurn B. A., Chantler S. M. Production of reagent antibodies. Methods Enzymol. 1980;70(A):104–142. doi: 10.1016/s0076-6879(80)70044-7. [DOI] [PubMed] [Google Scholar]
  12. Jacobson G. R., Rosenbusch J. P. ATP binding to a protease-resistant core of actin. Proc Natl Acad Sci U S A. 1976 Aug;73(8):2742–2746. doi: 10.1073/pnas.73.8.2742. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Konigsberg I. R. Diffusion-mediated control of myoblast fusion. Dev Biol. 1971 Sep;26(1):133–152. doi: 10.1016/0012-1606(71)90113-8. [DOI] [PubMed] [Google Scholar]
  14. Korn E. D. Actin polymerization and its regulation by proteins from nonmuscle cells. Physiol Rev. 1982 Apr;62(2):672–737. doi: 10.1152/physrev.1982.62.2.672. [DOI] [PubMed] [Google Scholar]
  15. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  16. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  17. Lerner R. A., Green N., Alexander H., Liu F. T., Sutcliffe J. G., Shinnick T. M. Chemically synthesized peptides predicted from the nucleotide sequence of the hepatitis B virus genome elicit antibodies reactive with the native envelope protein of Dane particles. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3403–3407. doi: 10.1073/pnas.78.6.3403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Linkhart T. A., Clegg C. H., Hauschika S. D. Myogenic differentiation in permanent clonal mouse myoblast cell lines: regulation by macromolecular growth factors in the culture medium. Dev Biol. 1981 Aug;86(1):19–30. doi: 10.1016/0012-1606(81)90311-0. [DOI] [PubMed] [Google Scholar]
  19. Lu R. C., Elzinga M. Partial amino acid sequence of brain actin and its homology with muscle actin. Biochemistry. 1977 Dec 27;16(26):5801–5806. doi: 10.1021/bi00645a025. [DOI] [PubMed] [Google Scholar]
  20. Pardee J. D., Bamburg J. R. Actin from embryonic chick brain. Isolation in high yield and comparison of biochemical properties with chicken muscle actin. Biochemistry. 1979 May 29;18(11):2245–2252. doi: 10.1021/bi00578a017. [DOI] [PubMed] [Google Scholar]
  21. Pollack R., Osborn M., Weber K. Patterns of organization of actin and myosin in normal and transformed cultured cells. Proc Natl Acad Sci U S A. 1975 Mar;72(3):994–998. doi: 10.1073/pnas.72.3.994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Rubenstein P. A., Spudich J. A. Actin microheterogeneity in chick embryo fibroblasts. Proc Natl Acad Sci U S A. 1977 Jan;74(1):120–123. doi: 10.1073/pnas.74.1.120. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Spudich J. A., Watt S. The regulation of rabbit skeletal muscle contraction. I. Biochemical studies of the interaction of the tropomyosin-troponin complex with actin and the proteolytic fragments of myosin. J Biol Chem. 1971 Aug 10;246(15):4866–4871. [PubMed] [Google Scholar]
  24. Storti R. V., Horovitch S. J., Scott M. P., Rich A., Pardue M. L. Myogenesis in primary cell cultures from Drosophila melanogaster: protein synthesis and actin heterogeneity during development. Cell. 1978 Apr;13(4):589–598. doi: 10.1016/0092-8674(78)90210-6. [DOI] [PubMed] [Google Scholar]
  25. Sutcliffe J. G., Shinnick T. M., Green N., Liu F. T., Niman H. L., Lerner R. A. Chemical synthesis of a polypeptide predicted from nucleotide sequence allows detection of a new retroviral gene product. Nature. 1980 Oct 30;287(5785):801–805. doi: 10.1038/287801a0. [DOI] [PubMed] [Google Scholar]
  26. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Vandekerckhove J., Leavitt J., Kakunaga T., Weber K. Coexpression of a mutant beta-actin and the two normal beta- and gamma-cytoplasmic actins in a stably transformed human cell line. Cell. 1980 Dec;22(3):893–899. doi: 10.1016/0092-8674(80)90566-8. [DOI] [PubMed] [Google Scholar]
  28. Vandekerckhove J., Weber K. Actin amino-acid sequences. Comparison of actins from calf thymus, bovine brain, and SV40-transformed mouse 3T3 cells with rabbit skeletal muscle actin. Eur J Biochem. 1978 Oct 16;90(3):451–462. doi: 10.1111/j.1432-1033.1978.tb12624.x. [DOI] [PubMed] [Google Scholar]
  29. Vandekerckhove J., Weber K. At least six different actins are expressed in a higher mammal: an analysis based on the amino acid sequence of the amino-terminal tryptic peptide. J Mol Biol. 1978 Dec 25;126(4):783–802. doi: 10.1016/0022-2836(78)90020-7. [DOI] [PubMed] [Google Scholar]
  30. Vandekerckhove J., Weber K. Mammalian cytoplasmic actins are the products of at least two genes and differ in primary structure in at least 25 identified positions from skeletal muscle actins. Proc Natl Acad Sci U S A. 1978 Mar;75(3):1106–1110. doi: 10.1073/pnas.75.3.1106. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Vandekerckhove J., Weber K. The amino acid sequence of actin from chicken skeletal muscle actin and chicken gizzard smooth muscle actin. FEBS Lett. 1979 Jun 15;102(2):219–222. doi: 10.1016/0014-5793(79)80004-6. [DOI] [PubMed] [Google Scholar]
  32. Vandekerckhove J., Weber K. The complete amino acid sequence of actins from bovine aorta, bovine heart, bovine fast skeletal muscle, and rabbit slow skeletal muscle. A protein-chemical analysis of muscle actin differentiation. Differentiation. 1979;14(3):123–133. doi: 10.1111/j.1432-0436.1979.tb01021.x. [DOI] [PubMed] [Google Scholar]
  33. Walter G., Hutchinson M. A., Hunter T., Eckhart W. Antibodies specific for the polyoma virus middle-size tumor antigen. Proc Natl Acad Sci U S A. 1981 Aug;78(8):4882–4886. doi: 10.1073/pnas.78.8.4882. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Walter G., Hutchinson M. A., Hunter T., Eckhart W. Purification of polyoma virus medium-size tumor antigen by immunoaffinity chromatography. Proc Natl Acad Sci U S A. 1982 Jul;79(13):4025–4029. doi: 10.1073/pnas.79.13.4025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Walter G., Scheidtmann K. H., Carbone A., Laudano A. P., Doolittle R. F. Antibodies specific for the carboxy- and amino-terminal regions of simian virus 40 large tumor antigen. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5197–5200. doi: 10.1073/pnas.77.9.5197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Whalen R. G., Butler-Browne G. S., Gros F. Protein synthesis and actin heterogeneity in calf muscle cells in culture. Proc Natl Acad Sci U S A. 1976 Jun;73(6):2018–2022. doi: 10.1073/pnas.73.6.2018. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Yaffe D. Developmental changes preceding cell fusion during muscle differentiation in vitro. Exp Cell Res. 1971 May;66(1):33–48. doi: 10.1016/s0014-4827(71)80008-3. [DOI] [PubMed] [Google Scholar]

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