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Journal of Solid-Phase Biochemistry

, Volume 1, Issue 4, pp 307–317 | Cite as

Affinity chromatography ofd-lactate dehydrogenases fromLimulus polyphemus (horseshoe crab) andHaliotus sp. (abalone) voluntary muscle on 8-(6-aminohexyl)-amino-adenine nucleotide-sepharose

  • George L. Long
  • Elizabeth L. Moore
Article
  • 23 Downloads

Abstract

An improved method of purification employing sequential isoenzyme elution on DEAE-cellulose at pH 6.5 and biologically specific elution with the reduced NAD-pyruvate adduct from 8-(6-aminohexyl)-amino-NAD⊕-Sepharose is described forLimulus (horseshoe crab) muscle D- lactate dehydrogenase. The protein is judged as being at least 98%pure by its constant specific activity in the terminal purification steps, a molar extinction coefficient (280nm) identical with that previously reported for the purified enzyme, and its protein and enzyme electrophoretic patterns on starch and polyacrylamide gel electrophoresis at three pHs. The binding of ammonium sulfate fractionated D- lactate dehydrogenase from crude cell- free Limulus andHaliotus (abalone) muscle homogenates to 8-(6-aminohexyl)-amino-AMP-and 8-(6-aminohexyl)-amino-NAD®-Sepharose columns is demonstrated. A comparison of the binding properties of these two enzymes with those of vertebrate L-lactate dehydrogenases suggests that they may be significantly different in terms of their binding sites for the adenine portion of the coenzyme.

Keywords

Lactate Dehydrogenase Ammonium Sulfate Horseshoe Crab Sepharose Column Disodium EDTA 
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.

References

  1. 1.
    Pesce, A., McKay, R. H., Stolzenbach, F., Cahn, R. D., andKaplan, N. O. (1964) J. Biol. Chem. 239: 1753.Google Scholar
  2. 2.
    Fondy, T. P., Pesce, A., Freedberg, I., Stolzenbach, F., andKaplan, N. O. (1964) Biochemistry 3: 522.CrossRefGoogle Scholar
  3. 3.
    Pesce, A., Fondy, T. P., Stolzenbach, F., Castillo, F., andKaplan, N. O. (1967) J. Biol. Chem. 242: 2151.Google Scholar
  4. 4.
    Fondy, T. P., Everse, J., Driscoll, G. A., Castillo, F., Stolzenbach, F. E., andKaplan, N. O. (1965) J. Biol. Chem. 240: 4219.Google Scholar
  5. 5.
    Kaloustian, H. D., Stolzenbach, F. E., Everse, J., andKaplan, N. O. (1969) J. Biol. Chem. 244: 2891.Google Scholar
  6. 6.
    Kaloustian, H. D., andKaplan, N. O. (1969) J. Biol. Chem. 244: 2902.Google Scholar
  7. 7.
    Eichner, R. D., andKaplan, N. O. (1977)Arch. Biochem. Biophys., in press.Google Scholar
  8. 8.
    Long, G. L., andKaplan, N. O. (1968) Science 162: 685.CrossRefGoogle Scholar
  9. 9.
    Long, G. L., andKaplan, N. O. (1973) Arch. Biochem. Biophys. 154: 696.CrossRefGoogle Scholar
  10. 10.
    Long, G. L., andKaplan, N. O. (1973) Arch. Biochem. Biophys. 154: 711.CrossRefGoogle Scholar
  11. 11.
    Long, G. L. (1975) “D-Lactate Dehydrogenase from the Horseshoe Crab,”In Methods in Enzymology, Vol. 41,Wood, W. A. (ed.), Academic Press, New York, pp. 313–318.Google Scholar
  12. 12.
    Lee, C.-Y., Lappi, D. A., Wermuth, B., Everse, J., andKaplan, N. O. (1974) Arch. Biochem. Biophys. 163: 561.CrossRefGoogle Scholar
  13. 13.
    Rossmann, M. G. (1975) “Evolutionary and Structural Relationships among Dehydrogenases,”In The Enzymes, Vol. 11,Boyer, P. D. (ed.), Academic Press, New York, pp. 62–102.Google Scholar
  14. 14.
    Lee, C.-Y., andKaplan, N. O. (1975) Arch. Biochem. Biophys. 168: 665.CrossRefGoogle Scholar
  15. 15.
    Fine, I. H., andCostello, L. (1963) “The Use of Starch Electrophoresis in Dehydrogenase Studies,”In Methods in Enzymology, Vol. 6,Colowick, S. P., andKaplan, N. O. (eds.), Academic Press, New York, pp. 958–972.CrossRefGoogle Scholar
  16. 16.
    Davis, B. J. (1964) Ann. N. Y. Acad. Sci. 121: 404.CrossRefGoogle Scholar
  17. 17.
    Gabriel, O. (1971) “Analytical Disc Gel Electrophoresis,”In Methods in Enzymology, Vol. 22,Jakoby, W. B. (ed.), Academic Press, New York, pp. 565–578.Google Scholar
  18. 18.
    Everse, J., Zoll, E.C., Kahan, L., andKaplan, N. O. (1971) Bioorg. Chem. 1: 207.CrossRefGoogle Scholar
  19. 19.
    Lee, C.-Y., andKaplan, N. O. (1976) J. Macromol. Sci. Chem. A10:15.CrossRefGoogle Scholar
  20. 20.
    Dixon, J. E. Personal communication (manuscript in preparation).Google Scholar

Copyright information

© Humana Press Inc. 1977

Authors and Affiliations

  • George L. Long
    • 1
  • Elizabeth L. Moore
    • 1
  1. 1.Department of ChemistryPomona CollegeClaremont

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