Isolation and some properties of 6-phosphogluconate dehydrogenase from Bacillus stearothermophilus

https://doi.org/10.1016/0005-2744(74)90148-XGet rights and content

Abstract

A purification procedure is described for 6-phosphogluconate dehdyrogenase (6-phospho-d-gluconate:NADP oxidoreductase (decarboxylating), EC 1.1.1.44) of Bacillus stearothermophilus (NCA 1503). A 1200-fold purification was obtained with a 15% yield. The purified enzyme has an approximate mol. wt of 101 000 as estimated by sucrose density gradient centrifugation and consists of subunits of a mol. wt of 51 000. Sulfhydryl group(s) are essential for enzymatic activity as indicated by p-chloromercuribenzoate and Ellman's reagent inactivation. Mg2+-activate the enzyme at a low concentration (0.01–0.04 M), whereas they inhibit at a higher concentration.

The optimum of the activity was found at about pH 8, with a Km value at 43 °C of 2.5·10−5 M for NADP and 2.0·10−5 M for 6-phosphogluconic acid. The enzyme is stable at 60 °C, whereas it is inactivated at higher temperatures, with a denaturation half-time of 80 min at 70 °C and 3 min at 80 °C. The enzyme shows a broken Arrhennius plot, with two straight lines meeting at about 50 °C.

References (32)

  • W.A. Scott et al.

    J. Biol. Chem.

    (1973)
  • S. Pontremoli et al.

    J. Biol. Chem.

    (1961)
  • M. Rippa et al.

    J. Biol. Chem.

    (1966)
  • M. Rippa et al.

    Arch. Biochem. Biophys.

    (1971)
  • M. Rippa et al.

    Arch. Biochem. Biophys.

    (1972)
  • K. Suzuki et al.

    FEBS Lett.

    (1971)
  • S. Sugimoto et al.

    Biochim. Biophys. Acta

    (1971)
  • W.H. Murphey et al.

    J. Biol. Chem.

    (1967)
  • K. Weber et al.

    J. Biol. Chem.

    (1969)
  • R.G. Martin et al.

    J. Biol. Chem.

    (1961)
  • G. Ashwell et al.

    J. Biol. Chem.

    (1957)
  • H.K. Kuramitsu

    J. Biol. Chem.

    (1970)
  • L.A. Fahien et al.

    Archiv. Biochem. Biophys.

    (1969)
  • R.H. Villet et al.

    Biochem. J.

    (1969)
  • R.H. Villet et al.

    Biochem. J.

    (1967)
  • R.H. Villet et al.

    Eur. J. Biochem.

    (1972)
  • Cited by (28)

    • Brain protection by rapeseed oil in magnesium-deficient mice

      2011, Prostaglandins Leukotrienes and Essential Fatty Acids
      Citation Excerpt :

      In a previous work, anti-NMDA properties of magnesium were shown to take place in vivo [22]. On the other hand, magnesium is a cofactor for the enzymes in glutathione biosynthesis (γ-glutamylcysteine synthetase and glutathione synthetase) [29,30] and NADPH-producing pentose phosphate pathway (6-phosphogluconate dehydrogenase and transketolase) [31,32], explaining why magnesium deficiency may affect reduced glutathione biosynthesis and recycling. In normal diet conditions, the physiological cellular loss of reduced glutathione is counterbalanced by intracellular glutathione biosynthesis.

    • Physiology of Thermophilic Bacteria

      1979, Advances in Microbial Physiology
    View all citing articles on Scopus

    A preliminary report of this work has been presented at the 9th Int. Congress of Biochemistry 1–7 July, Stockholm, Abst. of Commun., 2 K 12.

    View full text