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. 1985 Apr;48(1):198–203. doi: 10.1128/iai.48.1.198-203.1985

Expression of protective and cardiac tissue cross-reactive epitopes of type 5 streptococcal M protein in Escherichia coli.

T P Poirier, M A Kehoe, J B Dale, K N Timmis, E H Beachey
PMCID: PMC261935  PMID: 2579908

Abstract

The immunochemical properties of type 5 M protein antigens that were expressed in Escherichia coli K-12 by recombinant lambda bacteriophages isolated from a gene bank of serotype 5 Streptococcus pyogenes have been analyzed in detail. M proteins from partially purified bacteriophage lysates displayed precipitin lines of identity with a purified peptic extract of type 5 M protein (pep M5) in immunodiffusion assays. Immunoblot analyses of the M protein-positive lysates demonstrated that the cloned M protein component resided in five polypeptides with relative molecular weights of 57,900 (57.9K), 55.4K, 52.9K, 40.0K, and 32.6K. The hybrid lambda phage (lambda M5)-produced M protein contained immunoprotective epitopes; lambda M5 protein inhibited opsonization of type 5 streptococci by pep M5 antibodies, and antiserum raised against lambda M5 lysates opsonized type 5 streptococci. Each of the five antigenic polypeptides of the recombinant phage M protein also shared epitopes with human heart tissue, as demonstrated by the reactivity of immunoblots of lambda M5 antigens separated on sodium dodecyl sulfate gels with anti-pep M5 antibodies absorbed to and eluted from human heart sarcolemmal membranes. Moreover, antiserum raised against the lambda M5 lysates reacted with sarcolemmal membrane proteins with relative molecular weights of 200K, 59K, 55K, 53K, and 27K as determined by immunoblot analyses. These results demonstrate that the structural gene coding for type 5 streptococcal M protein which was inserted into lambda DNA expresses immunoprotective epitopes, some of which are shared with human heart tissue.

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Selected References

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

  1. Beachey E. H., Campbell G. L., Ofek I. Peptic digestion of streptococcal M protein. II. Extraction of M antigen from group A streptococci with pepsin. Infect Immun. 1974 May;9(5):891–896. doi: 10.1128/iai.9.5.891-896.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Beachey E. H., Seyer J. M., Kang A. H. Primary structure of protective antigens of type 24 streptococcal M protein. J Biol Chem. 1980 Jul 10;255(13):6284–6289. [PubMed] [Google Scholar]
  3. Beachey E. H., Stollerman G. H., Chiang E. Y., Chiang T. M., Seyer J. M., Kang A. H. Purification and properties of M protein extracted from group A streptococci with pepsin: covalent structure of the amino terminal region of type 24 M antigen. J Exp Med. 1977 Jun 1;145(6):1469–1483. doi: 10.1084/jem.145.6.1469. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Beachey E. H., Stollerman G. H., Johnson R. H., Ofek I., Bisno A. L. Human immune response to immunization with a structurally defined polypeptide fragment of streptococcal M protein. J Exp Med. 1979 Oct 1;150(4):862–877. doi: 10.1084/jem.150.4.862. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Dale J. B., Beachey E. H. Protective antigenic determinant of streptococcal M protein shared with sarcolemmal membrane protein of human heart. J Exp Med. 1982 Oct 1;156(4):1165–1176. doi: 10.1084/jem.156.4.1165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dale J. B., Beachey E. H. Unique and common protective epitopes among different serotypes of group A streptococcal M proteins defined with hybridoma antibodies. Infect Immun. 1984 Oct;46(1):267–269. doi: 10.1128/iai.46.1.267-269.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dale J. B., Ofek I., Beachey E. H. Heterogeneity of type-specific and cross-reactive antigenic determinants within a single M protein of group A streptococci. J Exp Med. 1980 May 1;151(5):1026–1038. doi: 10.1084/jem.151.5.1026. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Engvall E., Perlmann P. Enzyme-linked immunosorbent assay, Elisa. 3. Quantitation of specific antibodies by enzyme-labeled anti-immunoglobulin in antigen-coated tubes. J Immunol. 1972 Jul;109(1):129–135. [PubMed] [Google Scholar]
  9. Fischetti V. A., Jones K. F., Manjula B. N., Scott J. R. Streptococcal M6 protein expressed in Escherichia coli. Localization, purification, and comparison with streptococcal-derived M protein. J Exp Med. 1984 Apr 1;159(4):1083–1095. doi: 10.1084/jem.159.4.1083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fischetti V. A. Streptococcal M protein extracted by nonionic detergent. II. Analysis of the antibody response to the multiple antigenic determinants of the M-protein molecule. J Exp Med. 1977 Oct 1;146(4):1108–1123. doi: 10.1084/jem.146.4.1108. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fox E. N. M proteins of group A streptococci. Bacteriol Rev. 1974 Mar;38(1):57–86. doi: 10.1128/br.38.1.57-86.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. KAPLAN M. H., MEYESERIAN M. An immunological cross-reaction between group-A streptococcal cells and human heart tissue. Lancet. 1962 Apr 7;1(7232):706–710. doi: 10.1016/s0140-6736(62)91653-7. [DOI] [PubMed] [Google Scholar]
  13. Kehoe M. A., Poirier T. P., Beachey E. H., Timmis K. N. Cloning and genetic analysis of serotype 5 M protein determinant of group A streptococci: evidence for multiple copies of the M5 determinant in the Streptococcus pyogenes genome. Infect Immun. 1985 Apr;48(1):190–197. doi: 10.1128/iai.48.1.190-197.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. LANCEFIELD R. C. Current knowledge of type-specific M antigens of group A streptococci. J Immunol. 1962 Sep;89:307–313. [PubMed] [Google Scholar]
  15. 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]
  16. Manjula B. N., Acharya A. S., Mische S. M., Fairwell T., Fischetti V. A. The complete amino acid sequence of a biologically active 197-residue fragment of M protein isolated from type 5 group A streptococci. J Biol Chem. 1984 Mar 25;259(6):3686–3693. [PubMed] [Google Scholar]
  17. OUCHTERLONY O. Diffusion-in-gel methods for immunological analysis. II. Prog Allergy. 1962;6:30–154. doi: 10.1159/000313795. [DOI] [PubMed] [Google Scholar]
  18. Phillips G. N., Jr, Flicker P. F., Cohen C., Manjula B. N., Fischetti V. A. Streptococcal M protein: alpha-helical coiled-coil structure and arrangement on the cell surface. Proc Natl Acad Sci U S A. 1981 Aug;78(8):4689–4693. doi: 10.1073/pnas.78.8.4689. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Scott J. R., Fischetti V. A. Expression of streptococcal M protein in Escherichia coli. Science. 1983 Aug 19;221(4612):758–760. doi: 10.1126/science.6192499. [DOI] [PubMed] [Google Scholar]
  20. Seyer J. M., Kang A. H., Beachey E. H. Primary structural similarities between types 5 and 24 M proteins of Streptococcus pyogenes. Biochem Biophys Res Commun. 1980 Jan 29;92(2):546–553. doi: 10.1016/0006-291x(80)90368-x. [DOI] [PubMed] [Google Scholar]
  21. Spanier J. G., Jones S. J., Cleary P. Small DNA deletions creating avirulence in Streptococcus pyogenes. Science. 1984 Aug 31;225(4665):935–938. doi: 10.1126/science.6089334. [DOI] [PubMed] [Google Scholar]
  22. 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]
  23. Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]
  24. Zabriskie J. B., Freimer E. H. An immunological relationship between the group. A streptococcus and mammalian muscle. J Exp Med. 1966 Oct 1;124(4):661–678. doi: 10.1084/jem.124.4.661. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. van de Rijn I., Fischetti V. A. Immunochemical analysis of intact M protein secreted from cell wall-less streptococci. Infect Immun. 1981 Apr;32(1):86–91. doi: 10.1128/iai.32.1.86-91.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. van de Rijn I., Zabriskie J. B., McCarty M. Group A streptococcal antigens cross-reactive with myocardium. Purification of heart-reactive antibody and isolation and characterization of the streptococcal antigen. J Exp Med. 1977 Aug 1;146(2):579–599. doi: 10.1084/jem.146.2.579. [DOI] [PMC free article] [PubMed] [Google Scholar]

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