Biochemical characterization of two glutamate dehydrogenases with different cofactor specificities from a hyperthermophilic archaeon Pyrobaculum calidifontis

Extremophiles. 2013 May;17(3):379-89. doi: 10.1007/s00792-013-0527-7. Epub 2013 Mar 19.


Two putative glutamate dehydrogenase (GDH) genes (pcal_1031 and pcal_1606) were found in a sulfur-dependent hyperthermophilic archaeon, Pyrobaculum calidifontis. The two genes were then expressed in Escherichia coli, and both of the recombinant gene products showed GDH activity. The two enzymes were then purified to homogeneity and characterized in detail. Although both purified GDHs had a hexameric structure and neither exhibited allosteric regulation, they showed different coenzyme specificities: one was specific for NAD(+), the other for NADP(+) and different heat activation mechanisms. In addition, there was little difference in the kinetic constants, optimal temperature, thermal stability, optimal pH and pH stability between the two enzymes. The overall sequence identity between the two proteins was very high (81%), but was not high in the region recognizing the 2' position of the adenine ribose moiety, which is responsible for coenzyme specificity. This is the first report on the identification of two GDHs with different coenzyme specificities from a single hyperthermophilic archaeon and the definition of their basic in vitro properties.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation
  • Amino Acid Sequence
  • Archaeal Proteins / chemistry*
  • Archaeal Proteins / genetics
  • Archaeal Proteins / metabolism
  • Glutamate Dehydrogenase / chemistry*
  • Glutamate Dehydrogenase / genetics
  • Glutamate Dehydrogenase / metabolism
  • Hydrogen-Ion Concentration
  • Kinetics
  • Molecular Sequence Data
  • NAD / metabolism
  • NADP / metabolism
  • Phylogeny
  • Protein Multimerization
  • Protein Stability
  • Pyrobaculum / enzymology*
  • Substrate Specificity
  • Temperature


  • Archaeal Proteins
  • NAD
  • NADP
  • Glutamate Dehydrogenase