Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1967 Feb;102(2):423–431. doi: 10.1042/bj1020423

Purification and properties of crystalline 3-hydroxybutyrate dehydrogenase from Rhodopseudomonas spheroides

H U Bergmeyer 1, K Gawehn 1, H Klotzsch 1, H A Krebs 1, D H Williamson 1
PMCID: PMC1270263  PMID: 4291491

Abstract

1. The purification and crystallization of 3-hydroxybutyrate dehydrogenase from extracts of Rhodopseudomonas spheroides is described. 2. The molecular weight was calculated to be 85000 by sedimentation equilibrium. 3. Although the enzyme is stable at 0–4°, dilute solutions are rapidly inactivated at 37°; NADH2 or Ca2+ ions prevent this inactivation. 4. The enzyme is extremely sensitive to mercurials, but can be protected by NADH2 or Ca2+ ions. 5. From studies on p-hydroxymercuribenzoate binding it is estimated that the enzyme contains 5–6 moles of rapidly reacting thiol groups/mole. 6. d-Lactate and dl-2-hydroxybutyrate are competitive inhibitors of d-3-hydroxybutyrate oxidation. 7. The properties of the crystalline enzyme are compared with those of 3-hydroxybutyrate dehydrogenase preparations from other sources.

Full text

PDF
424

Images in this article

424Fig. 1.
on p.424

Selected References

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

  1. BERGMEYER H. U., BERNT E. ENZYMATISCHE BESTIMMUNG VON KETON-KOERPERN IM BLUT. Enzymol Biol Clin (Basel) 1965;19:65–76. [PubMed] [Google Scholar]
  2. BERRY M. N. SOME INHIBITORS OF D-3-HYDROXYBUTYRATE DEHYDROGENASE. Biochim Biophys Acta. 1964 Oct 23;92:156–159. doi: 10.1016/0926-6569(64)90282-2. [DOI] [PubMed] [Google Scholar]
  3. DIXON M. The determination of enzyme inhibitor constants. Biochem J. 1953 Aug;55(1):170–171. doi: 10.1042/bj0550170. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Delafield F. P., Cooksey K. E., Doudoroff M. beta-Hydroxybutyric dehydrogenase and dimer hydrolase of Pseudomonas lemoignei. J Biol Chem. 1965 Oct;240(10):4023–4028. [PubMed] [Google Scholar]
  5. Edson N. L. Ketogenesis-antiketogenesis: The influence of ammonium chloride on ketone-body formation in liver. Biochem J. 1935 Sep;29(9):2082–2094. doi: 10.1042/bj0292082. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. HALL L. M. Preparation of crystalline lithium acetoacetate. Anal Biochem. 1962 Jan;3:75–80. doi: 10.1016/0003-2697(62)90046-5. [DOI] [PubMed] [Google Scholar]
  7. OKUNUKI K. Denaturation and inactivation of enzyme proteins. Adv Enzymol Relat Subj Biochem. 1961;23:29–82. doi: 10.1002/9780470122686.ch2. [DOI] [PubMed] [Google Scholar]
  8. SEKUZU I., JURTSHUK P., Jr, GREEN D. E. Studies on the electron transfer system. LI. Isolation and characterization of the D-(--)-beta-hydroxybutyric apodehydrogenase from beef heart mitochondria. J Biol Chem. 1963 Mar;238:975–982. [PubMed] [Google Scholar]
  9. SHUSTER C. W., DOUDOROFF M. A cold-sensitive D(-) beta-hydroxybutyric acid dehydrogenase from Rhodospirillum rubrum. J Biol Chem. 1962 Feb;237:603–607. [PubMed] [Google Scholar]
  10. WILLIAMSON D. H., MELLANBY J., KREBS H. A. Enzymic determination of D(-)-beta-hydroxybutyric acid and acetoacetic acid in blood. Biochem J. 1962 Jan;82:90–96. doi: 10.1042/bj0820090. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. WISE J. B., LEHNINGER A. L. The stability of mitochondrial D-beta-hydroxybutyric dehydrogenase and its relationship to the respiratory chain. J Biol Chem. 1962 Apr;237:1363–1370. [PubMed] [Google Scholar]
  12. YPHANSTIS D. A. Rapid determination of molecular weights of peptides and preteins. Ann N Y Acad Sci. 1960 Aug 31;88:586–601. doi: 10.1111/j.1749-6632.1960.tb20055.x. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES