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Purification and Characterization of Hypoxanthine-Guanine Phosphoribosyl Transferase from Cultured HTC Cells

  • Vassilis I. Zannis
  • Lorraine J. Gudas
  • Deborah Doyle
  • David W. MartinJr.
Chapter
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Part of the Advances in Experimental Medicine and Biology book series

Abstract

Hypoxanthine-guanine phosphoribosyl transferase (HGPRTase.) from |35S|-methionyl-labelled rat hepatoma (HTC) cells has been purified more than 600-fold to near homogeneity by making use of conventional and affinity chromatographic techniques (Table 1). The conventional purification procedure utilizes a pH 5.0 precipitation, an ammonium sulfate precipitation, DEAE-cellulose chromatography, and heating at 80–85°according to the method of Olsen and Milman, 1974. Using this procedure, a partially pure enzyme preparation is obtained. Affinity chromatography according to Hughes et al., 1975, can be used either before or after the DEAE step to yield pure enzyme.

Keywords

Crude Extract Pure Enzyme Preparation Enlarge Portion Potassium Phosphate Buffer 60mM Major Isoenzyme 
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.

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References

  1. 1.
    Chasin, L.A. and Urlaub, G. (1976) Somatic Cell Genetics 2, 453.PubMedCrossRefGoogle Scholar
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    Graf, L.H., McRoberts, J.A., Harrison, T. and Martin, D.W. (1976) J. Cell. Physiol. 80, 331.CrossRefGoogle Scholar
  3. 3.
    Hughes, S.H., Wahl, M. and Capecchi, M.R. (1975) J. Biol. Chem. 250, 120.PubMedGoogle Scholar
  4. 4.
    O’Farrell, P.H. (1975) J. Biol. Chem. 250, 4007.PubMedGoogle Scholar
  5. 5.
    Olsen, A.S. and Milman, G. (1974) J. Biol. Chem. 249, 4030.PubMedGoogle Scholar

Copyright information

© Plenum Press, New York 1977

Authors and Affiliations

  • Vassilis I. Zannis
    • 1
  • Lorraine J. Gudas
    • 1
  • Deborah Doyle
    • 1
  • David W. MartinJr.
    • 1
  1. 1.Departments of Medicine and Biochemistry and BiophysicsUniversity of California, San FranciscoSan FranciscoUSA

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