Skip to main content
PMC home page
The Journal of Experimental Medicine logoLink to The Journal of Experimental Medicine
. 1967 May 1;125(5):893–920. doi: 10.1084/jem.125.5.893

PHYLOGENY OF IMMUNOGLOBULIN STRUCTURE AND FUNCTION

I. IMMUNOGLOBULINS OF THE LEMON SHARK

L W Clem 1, P A Small Jr 1
PMCID: PMC2138209  PMID: 4164694

Abstract

Lemon sharks immunized with bovine serum albumin produced two molecular forms of antibodies detectable by passive hemagglutination of antigen-coated, tanned sheep erythrocytes. Throughout the course of immunization 2-ME-sensitive antibody was associated with a 19S immunoglobulin fraction (4–5 mg/ml serum) while late in the course of immunization antibody was found also associated with a 7S immunoglobulin fraction (7–8 mg/ml serum). No evidence for any anamnestic response was found in these animals. Naturally occurring hemagglutinins for sheep erythrocytes were found to be 2-ME-sensitive and present in the 19S immunoglobulin fraction. These immunoglobulin fractions were readily purified by DEAE-cellulose chromatography and Sephadex G-200 gel filtration. Both immunoglobulin molecules yielded equimolar amounts of H and L polypeptide chains when subjected to extensive reduction and alkylation followed by gel filtration in 5 M guanidine-HCl. Antigenically reactive H and L chains were obtained by partial reduction and alkylation followed by gel filtration in 1 M propionic acid. The 7S and 19S immunoglobulin H chains were indistinguishable by fingerprints of tryptic digests, disc electrophoretic patterns, antigenic properties, and mass (molecular weight ∼70,000), thus suggesting these two molecules to belong to the same immunoglobulin class. The shark 19S and 7S immunoglobulin L chains were indistinguishable from each other by similar criteria and were different from the H chains. These L chains exhibited the electrophoretic heterogeneity of their mammalian counterparts. The 7S (shark immunoglobulin) molecule was shown to have a molecular weight of ∼160,000 and to consist of 2H and 2L polypeptide chains (total mass ≅180,000). The 19S molecule was shown to have a molecular weight of 800,000–900,000; therefore, there were probably five 7S subunits per 19S molecule, comparable to mammalian γM. Other reasons for considering the 7S and the 19S lemon shark molecules to belong to a class of immunoglobulins comparable to the γM class of mammals are that they both have high carbohydrate contents, and H chains of mass similar to µ chains. The lemon shark serum proteins with electrophoretic mobilities comparable to gamma G of mammals were not related to the immunoglobulins of this species. These proteins had no antibody activity and had no antigenic or chemical similarity to either the H chains, the L chains, or the intact immunoglobulin molecules from the lemon shark.

Full Text

The Full Text of this article is available as a PDF (1.4 MB).

Selected References

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

  1. Andrews P. Estimation of the molecular weights of proteins by Sephadex gel-filtration. Biochem J. 1964 May;91(2):222–233. doi: 10.1042/bj0910222. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. BAUER D. C., MATHIES M. J., STAVITSKY A. B. Sequences of synthesis of gamma-1 macroglobulin and gamma-2 globulin antibodies during primary and secondary responses to proteins, salmonella antigens, and phage. J Exp Med. 1963 Jun 1;117:889–907. doi: 10.1084/jem.117.6.889. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. CLEM L. W., SIGEL M. M. COMPARATIVE IMMUNOCHEMICAL AND IMMUNOLOGICAL REACTIONS IN MARINE FISHES WITH SOLUBLE, VIRAL, AND BACTERIAL ANTIGENS. Fed Proc. 1963 Jul-Aug;22:1138–1144. [PubMed] [Google Scholar]
  4. COHEN S., PORTER R. B. STRUCTURE AND BIOLOGICAL ACTIVITY OF IMMUNOGLOBULINS. Adv Immunol. 1964;27:287–349. doi: 10.1016/s0065-2776(08)60710-5. [DOI] [PubMed] [Google Scholar]
  5. DEUTSCH H. F., MORTON J. I. Dissociation of human serum macroglobulins. Science. 1957 Mar 29;125(3248):600–601. doi: 10.1126/science.125.3248.600. [DOI] [PubMed] [Google Scholar]
  6. EDELMAN G. M., KUNKEL H. G., FRANKLIN E. C. Interaction of the rheumatoid factor with antigen-antibody complexes and aggregated gamma globulin. J Exp Med. 1958 Jul 1;108(1):105–120. doi: 10.1084/jem.108.1.105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. FLEISCHMAN J. B., PAIN R. H., PORTER R. R. Reduction of gamma-globulins. Arch Biochem Biophys. 1962 Sep;Suppl 1:174–180. [PubMed] [Google Scholar]
  8. Freeman M. J., Stavitsky A. B. Radioimmunoelectrophoretic study of rabbit anti-protein antibodies during the primary response. J Immunol. 1965 Dec;95(6):981–990. [PubMed] [Google Scholar]
  9. GOOD R. A., PAPERMASTER B. W. ONTOGENY AND PHYLOGENY OF ADAPTIVE IMMUNITY. Adv Immunol. 1964;27:1–115. doi: 10.1016/s0065-2776(08)60706-3. [DOI] [PubMed] [Google Scholar]
  10. GREENBURY C. L., MOORE D. H., NUNN L. A. REACTION OF 7S AND 19S COMPONENTS OF IMMUNE RABBIT ANTISERA WITH HUMAN GROUP A AND AB RED CELLS. Immunology. 1963 Sep;6:421–433. [PMC free article] [PubMed] [Google Scholar]
  11. GREY H. M. PHYLOGENY OF THE IMMUNE RESPONSE. STUDIES ON SOME PHYSICAL CHEMICAL AND SEROLOGIC CHARACTERISTICS OF ANTIBODY PRODUCED IN THE TURTLE. J Immunol. 1963 Dec;91:819–825. [PubMed] [Google Scholar]
  12. HVIDT A., JOHANSEN G., LINDERSTRØM LANG K., VASLOW F. Exchange of deuterium and 18O between water and other substances. I. Methods. C R Trav Lab Carlsberg Chim. 1954;29(9):129–157. [PubMed] [Google Scholar]
  13. Hilschmann N., Craig L. C. Amino acid sequence studies with Bence-Jones proteins. Proc Natl Acad Sci U S A. 1965 Jun;53(6):1403–1409. doi: 10.1073/pnas.53.6.1403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. KATZ A. M., DREYER W. J., ANFINSEN C. B. Peptide separation by two-dimensional chromatography and electrophoresis. J Biol Chem. 1959 Nov;234:2897–2900. [PubMed] [Google Scholar]
  15. LEVY H. B., SOBER H. A. A simple chromatographic method for preparation of gamma globulin. Proc Soc Exp Biol Med. 1960 Jan;103:250–252. doi: 10.3181/00379727-103-25476. [DOI] [PubMed] [Google Scholar]
  16. Lamm M. E., Small P. A., Jr Polypeptide chain structure of rabbit immunoglobulins. II. gamma-M-immunoglobulin. Biochemistry. 1966 Jan;5(1):267–276. doi: 10.1021/bi00865a035. [DOI] [PubMed] [Google Scholar]
  17. MILLER F., METZGER H. CHARACTERIZATION OF A HUMAN MACROGLOBULIN. I. THE MOLECULAR WEIGHT OF ITS SUBUNIT. J Biol Chem. 1965 Aug;240:3325–3333. [PubMed] [Google Scholar]
  18. Marchalonis J., Edelman G. M. Phylogenetic origins of antibody structure. I. Multichain structure of immunoglobulins in the smooth dogfish (Mustelus canis). J Exp Med. 1965 Sep 1;122(3):601–618. doi: 10.1084/jem.122.3.601. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Milstein C. Variations in amino-acid sequence near the disulphide bridges of Bence-Jones proteins. Nature. 1966 Jan 22;209(5021):370–373. doi: 10.1038/209370a0. [DOI] [PubMed] [Google Scholar]
  20. Onoue K., Yagi Y., Grossberg A. L., Pressman D. Number of binding sites of rabbit macroglobulin antibody and its subunits. Immunochemistry. 1965 Dec;2(4):401–415. doi: 10.1016/0019-2791(65)90039-x. [DOI] [PubMed] [Google Scholar]
  21. PAPERMASTER B. W., CONDIE R. M., FINSTAD J., GOOD R. A. EVOLUTION OF THE IMMUNE RESPONSE. I. THE PHYLOGENETIC DEVELOPMENT OF ADAPTIVE IMMUNOLOGIC RESPONSIVENESS IN VERTEBRATES. J Exp Med. 1964 Jan 1;119:105–130. doi: 10.1084/jem.119.1.105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. PORTER R. R. Separation and isolation of fractions of rabbit gamma-globulin containing the antibody and antigenic combining sites. Nature. 1958 Sep 6;182(4636):670–671. doi: 10.1038/182670a0. [DOI] [PubMed] [Google Scholar]
  23. Reisfeld R. A., Small P. A., Jr Electrophoretic heterogeneity of polypeptide chains of specific antibodies. Science. 1966 May 27;152(3726):1253–1255. doi: 10.1126/science.152.3726.1253. [DOI] [PubMed] [Google Scholar]
  24. SCHEIDEGGER J. J. Une micro-méthode de l'immuno-electrophorèse. Int Arch Allergy Appl Immunol. 1955;7(2):103–110. [PubMed] [Google Scholar]
  25. SMALL P. A., Jr, REISFELD R. A., DRAY S. PEPTIDE DIFFERENCES OF RABBIT GAMMA-G-IMMUNOGLOBULIN LIGHT CHAINS CONTROLLED BY ALLELIC GENES. J Mol Biol. 1965 Apr;11:713–721. doi: 10.1016/s0022-2836(65)80029-8. [DOI] [PubMed] [Google Scholar]
  26. STAVITSKY A. B. Micromethods for the study of proteins and antibodies. I. Procedure and general applications of hemagglutination and hemagglutination-inhibition reactions with tannic acid and protein-treated red blood cells. J Immunol. 1954 May;72(5):360–367. [PubMed] [Google Scholar]
  27. SVEHAG S. E., MANDEL B. THE FORMATION AND PROPERTIES OF POLIOVIRUS-NEUTRALIZING ANTIBODY. II. 19S AND 7S ANTIBODY FORMATION: DIFFERENCES IN ANTIGEN DOSE REQUIREMENT FOR SUSTAINED SYNTHESIS, ANAMNESIS, AND SENSITIVITY TO X-IRRADIATION. J Exp Med. 1964 Jan 1;119:21–39. doi: 10.1084/jem.119.1.21. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sigel M. M., Clem L. W. Antibody response of fish to viral antigens. Ann N Y Acad Sci. 1965 Aug 10;126(1):662–677. doi: 10.1111/j.1749-6632.1965.tb14312.x. [DOI] [PubMed] [Google Scholar]
  29. Small P. A., Jr, Lamm M. E. Polypeptide chain structure of rabbit immunoglobulins. I. gamma-G-immunoglobulin. Biochemistry. 1966 Jan;5(1):259–267. doi: 10.1021/bi00865a034. [DOI] [PubMed] [Google Scholar]
  30. Titani K., Whitley E., Jr, Avogardo L., Putnam F. W. Immunoglobulin structure: partial amino acid sequence of a Bence Jones protein. Science. 1965 Sep 3;149(3688):1090–1092. doi: 10.1126/science.149.3688.1090. [DOI] [PubMed] [Google Scholar]
  31. UHR J. W., FINKELSTEIN M. S. Antibody formation. IV. Formation of rapidly and slowly sedimenting antibodies and immunological memory to bacteriophage phi-X 174. J Exp Med. 1963 Mar 1;117:457–477. doi: 10.1084/jem.117.3.457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. UHR J. W., FINKELSTEIN S., FRANKLIN E. C. Antibody response to bacteriophage phi-X-174 in non-mammalian vertebrates. Proc Soc Exp Biol Med. 1962 Oct;111:13–15. doi: 10.3181/00379727-111-27691. [DOI] [PubMed] [Google Scholar]
  33. WINZLER R. J. Determination of serum glycoproteins. Methods Biochem Anal. 1955;2:279–311. doi: 10.1002/9780470110188.ch10. [DOI] [PubMed] [Google Scholar]
  34. YPHANTIS D. A. EQUILIBRIUM ULTRACENTRIFUGATION OF DILUTE SOLUTIONS. Biochemistry. 1964 Mar;3:297–317. doi: 10.1021/bi00891a003. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Experimental Medicine are provided here courtesy of The Rockefeller University Press

RESOURCES