title: Properties of glutamate dehydrogenase and its involvement in alanine production in a hyperthermophilic archaeon, Thermococcus profundus.
abstract: Thermococcus profundus, a hyperthermophilic archaeon, did not exhibit detectable glutamine synthetase activity, although the organism possessed an extraordinarily high level of glutamate dehydrogenase (GDH), the content of which reached over 10% of total soluble proteins. This GDH was purified to homogeneity. The enzyme had a molecular weight of 263,000 and was composed of six homogeneous subunits of molecular weight 43,000. The enzyme was extremely thermostable with a half life of 1 h at 90 degrees C. Circular dichroism (CD) spectra of the enzyme revealed gradual unfolding of alpha-helices upon exposure to increasing temperature. The enzyme reaction was strongly biased toward glutamate formation. T. profundus excreted L-alanine into the medium, and the concentration reached mM. High activity of alanine aminotransferase (AAT) was present in the cells, while no alanine dehydrogenase activity was detected. The alanine formation may be initiated by ammonia uptake by GDH followed by aminotransfer from glutamate to pyruvate by AAT.
authors.affiliation: Laboratory of Microbiology, Institute of Physical and Chemical Research (RIKEN), Saitama.
authors.forename: T
authors.initials: T
authors.lastname: Kobayashi
authors.forename: S
authors.initials: S
authors.lastname: Higuchi
authors.forename: K
authors.initials: K
authors.lastname: Kimura
authors.forename: T
authors.initials: T
authors.lastname: Kudo
authors.forename: K
authors.initials: K
authors.lastname: Horikoshi
mesh_terms: D000409:Alanine / Q000096:biosynthesis*; D000410:Alanine Transaminase / Q000378:metabolism; D000595:Amino Acid Sequence; D001105:Archaea / Q000201:enzymology*; D002942:Circular Dichroism; D004795:Enzyme Stability; D005969:Glutamate Dehydrogenase / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017386:Sequence Homology, Amino Acid; D013696:Temperature
chemical_list: D005969:Glutamate Dehydrogenase; D000410:Alanine Transaminase; D000409:Alanine