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. 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.

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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]

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