Skip to main content
PMC home page
Infection and Immunity logoLink to Infection and Immunity
. 1971 Jan;3(1):133–140. doi: 10.1128/iai.3.1.133-140.1971

Amino Acid Analysis and Molecular Weight Determination of Tetanus Toxin

Marlee J Holmes 1, Wayne L Ryan 1
PMCID: PMC416118  PMID: 16557929

Abstract

A procedure for the isolation of tetanus toxin was developed. The toxin was precipitated with ammonium sulfate and further purified by use of molecular sieve chromatography and preparative disc gel electrophoresis. The degree of purification resulting from these fractionation procedures was monitored by analytical poly-acrylamide gel electrophoresis, immunodiffusion, and immunoelectrophoresis, all three of which demonstrated a single component. There was a 50-fold increase in specific activity between the crude culture filtrate and the final purified product, which had an average specific activity of 300 × 106 mouse minimal lethal doses per mg of N. A molecular weight of 140,000 was estimated by gel filtration on Bio-Gel P-200. A complete amino acid analysis of tetanus toxin is presented.

Full text

PDF
134

Images in this article

136Image
on p.136
136Image
on p.136
136Image
on p.136

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. Bizzini B., Turpin A., Raynaud M. Production et purification de la toxine tétanique. Ann Inst Pasteur (Paris) 1969 May;116(5):686–712. [PubMed] [Google Scholar]
  3. Dawson D. J., Mauritzen C. M. Studies on tetanus toxin and toxoid. I. Isolation of tetanus toxin using DEAE-cellulose. Aust J Biol Sci. 1967 Feb;20(1):253–263. [PubMed] [Google Scholar]
  4. Dawson D. J., Mauritzen C. M. Studies on tetanus toxin and toxoid. II. Isolation and characterization of the tetanus toxin and toxoid. Aust J Biol Sci. 1968 Jun;21(3):559–568. doi: 10.1071/bi9680559. [DOI] [PubMed] [Google Scholar]
  5. HARDEGREE M. C. SEPARATION OF NEUROTOXIN AND HEMOLYSIN OF CLOSTRIDIUM TETANI. Proc Soc Exp Biol Med. 1965 Jun;119:405–408. doi: 10.3181/00379727-119-30195. [DOI] [PubMed] [Google Scholar]
  6. JOUBERT F. J., LARGIER J. F. Investigation of the physical properties of tetanus toxoids. Biochim Biophys Acta. 1956 May;20(2):407–408. doi: 10.1016/0006-3002(56)90313-4. [DOI] [PubMed] [Google Scholar]
  7. KORNGOLD L., VAN LEEUWEN G. The effect of the antigen's molecular weight on the curvature of the precipitin line in the Ouchterlony technic. J Immunol. 1957 Mar;78(3):172–177. [PubMed] [Google Scholar]
  8. Komorowska-Rycerz A., Brühl A., Krauze R. Fractionation of tetanus toxoid by gel filtration. Bull Acad Pol Sci Biol. 1966;14(2):81–84. [PubMed] [Google Scholar]
  9. LEVINE L., STONE J. L. The purification of tetanus toxoid by ammonium sulfate fractionation. J Immunol. 1951 Sep;67(3):235–242. [PubMed] [Google Scholar]
  10. 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]
  11. Latham W. C., Jenness C. P., Timperi R. J., Michelsen C. B., Zipilivan E. M., Edsall G., Ley H. L., Jr Purification and characterization of tetanus toxoid and toxin. I. Fractionation of tetanus toxoid by gel filtration. J Immunol. 1965 Sep;95(3):487–493. [PubMed] [Google Scholar]
  12. Murphy S. G., Miller K. D. Tetanus toxin and antigenic derivatives. I. Purification of the biologically active monomer. J Bacteriol. 1967 Sep;94(3):580–585. doi: 10.1128/jb.94.3.580-585.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Nielsen P. A., Ablondi F. B., Querry M. V., Gussoni C., Cooper M. S. Antigenic and immunogenic studies on purified tetanus toxoid. J Immunol. 1967 Jun;98(6):1248–1256. [PubMed] [Google Scholar]
  14. Pickett M. J., Hoeprich P. D., Germain R. O. Purification of High Titer Tetanus Toxin. J Bacteriol. 1945 May;49(5):515–516. doi: 10.1128/jb.49.5.515-516.1945. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Pillemer L., Wittler R., Grossberg D. B. The Isolation and Crystallization of Tetanal Toxin. Science. 1946 May 17;103(2681):615–616. doi: 10.1126/science.103.2681.615. [DOI] [PubMed] [Google Scholar]
  16. RAYNAUD M., TURPIN A., BIZZINI B. [Existence of tetanus toxin under several aggregation conditions]. Ann Inst Pasteur (Paris) 1960 Aug;99:167–172. [PubMed] [Google Scholar]
  17. Robb L. A. Separation of tetanus toxin and toxoid proteins by Sephadex gel. Can J Microbiol. 1967 Aug;13(8):1021–1025. doi: 10.1139/m67-136. [DOI] [PubMed] [Google Scholar]
  18. SHEFF M. F., PERRY M. B., ZACKS S. I. STUDIES ON TETANUS TOXIN. II. THE PREPARATION OF PURIFIED TETANUS TOXIN AND ITS FLUORESCENT LABELLING. Biochim Biophys Acta. 1965 Apr 12;100:215–234. [PubMed] [Google Scholar]
  19. Salenstedt C. R., Tirunarayanan M. O. Purification of tetanus toxin with the aid of Sephadex gels. Z Immunitatsforsch Allerg Klin Immunol. 1966 Feb;130(2):190–196. [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES