Advertisement

Purification of Terminal Deoxynucleotidyl Transferase from Pig Thymus: Identification of 42,000 and 57,000 Dalton Species

  • Tsuguhiro Kaneda
  • Saiko Kuroda
  • Osamu Koiwai
  • Shonen Yoshida
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 145)

Summary

Terminal deoxynucleotidyl transferase (TdT, EC 2.7.7.31) has been purified 4365-fold from pig thymus. It was further separated into two molecular forms of 57,000 and 45,000 dalton by Sephadex G-100 gel-filtration. The former sedimented at 4.2s through a sucrose gradient, while the latter, at 3.6s. By a SDS-polyacrylamide gel-electrophoresis, their molecular weights were estimated as 57,000 and 42,000 dalton, respectively. Thus each of the large and small pig TdT consists of a single polypeptide of 57,000 or 42,000 dalton and has no subunit structure. These two forms were indistinguishable in antigenicity by a neutralization assay with an anticalf TdT antibody. The enzymological properties of 42,000 dalton-TdT from pig thymus were very similar to those of calf TdT which has a two-subunits structure.

Keywords

Terminal Deoxynucleotidyl Transferase Single Polypeptide Subunit Structure High Molecular Weight Form 50mM Potassium Phosphate 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    F. J. Bollum, Oligodeoxyribonucleotideprimed reactions catalyzed by calf thymus polymerase, J.Biol. Chem. 237:1945 (1962).PubMedGoogle Scholar
  2. 2.
    L. M .S. Chang, Development of terminal deoxynucleotidyl transferase activity in embryonic calf thymus gland, Biochem. Biophys. Res. Commun. 44:124 (1971).PubMedCrossRefGoogle Scholar
  3. 3.
    M. S. Coleman, J. J. Hutton, P. D. Simone, and F. J. Bollum, Teminal deoxyribonucleotidyl transferase in human leukemia, Proc. Nat. Acad. Sci. USA 71:4404 (1974).PubMedCrossRefGoogle Scholar
  4. 4.
    R. D. Barr, P. S. Sarin, and S. M. Perry, Terminal transferase in human bone-marrow lymphocytes. Lancet 1:508 (1976).PubMedCrossRefGoogle Scholar
  5. 5.
    D. Baltimore, Is terminal deoxynucleotidyl transferase a somatic mutagen in lymphocytes ?, Nature 248:409 (1974).PubMedCrossRefGoogle Scholar
  6. 6.
    F. J. Bollum, “Karl August Forster Lecture,” Vol.14, pp.1–47, Franz Steiner Verlag, Wiesbaden (1975).Google Scholar
  7. 7.
    L. M. S. Chang and F. J. Bollum, Deoxynucleotide-polymerizing enzymes of calf thymus gland, J.Biol. Chem. 246:909 (1971)PubMedGoogle Scholar
  8. 8.
    F. J. Bollum and M. Brown, A high molecular weight form of terminal deoxynucleotidyl transferase, Nature 278:191 (1979).PubMedCrossRefGoogle Scholar
  9. 9.
    M. R. Deibel, Jr. and M. S. Coleman, Purification of a high molecular weight human terminal deoxynucleotidyl transferase, J.Biol. Chem. 254:8634 (1979).PubMedGoogle Scholar
  10. 10.
    A. Silverstone, L. Sun, O. N. Witte, and D. Baltimore, Biosynthesis of murine terminal deoxynucleotidyltransferase, J.Biol. Chem. 255:791 (1980)PubMedGoogle Scholar
  11. 11.
    R. C. Switzer, III, C. R. Merril, and S. Shifrin, A highly sensitive silver stain for detecting proteins and peptides in Polyacrylamide gels. Anal. Biochem. 98:231 (1979).PubMedCrossRefGoogle Scholar
  12. 12.
    U. K. Laemmli, Cleavage of structural proteins during the assembly of the head of bacteriophage T4, Nature 227:680 (1970)PubMedCrossRefGoogle Scholar
  13. 13.
    T. Kaneda, H. Mishima, Y. Hirota, and S. Yoshida, Immunological comparison of terminal deoxynucleotidyl transferases from various mammals in terms of calf thymus enzyme, J.Appl. Biochem. in press,Google Scholar
  14. 14.
    R. A. Dicioccio, K. Chadha, and B. I. S. Srivastava, Inhibition of herpes simplex virus-induced DNA polymerase, cellular DNA polymerase a, and virus production by aphidicolin, Biochim, Biophys,Acta 609:224 (1980).CrossRefGoogle Scholar
  15. 15.
    K. Kato, J. M. Goncalves, G. E. Houts, and F. J. Bollum, Deoxynucleo tide-polymerizing enzymes of calf thymus gland, J.Biol. Chem. 242:2780 (1967).PubMedGoogle Scholar

Copyright information

© Plenum Press, New York 1982

Authors and Affiliations

  • Tsuguhiro Kaneda
    • 1
  • Saiko Kuroda
    • 1
  • Osamu Koiwai
    • 2
  • Shonen Yoshida
    • 2
  1. 1.Blood Disease CenterNagoya National HospitalNaka-ku, Nagoya 460Japan
  2. 2.Department of BiochemistryInstitute for Developmental ResearchKasugai 480-03Japan

Personalised recommendations