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

Taisuke Wakamatsu, Chisato Higashi, Taketo Ohmori, Katsumi Doi, Toshihisa Ohshima

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)379-389
Number of pages11
JournalExtremophiles
Volume17
Issue number3
DOIs
Publication statusPublished - Mar 19 2013

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Pyrobaculum
Glutamate Dehydrogenase
Coenzymes
Archaea
Hot Temperature
Allosteric Regulation
Genes
Ribose
Adenine
Enzymes
NADP
Sulfur
NAD
Escherichia coli
Temperature
Proteins

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Molecular Medicine

Cite this

Biochemical characterization of two glutamate dehydrogenases with different cofactor specificities from a hyperthermophilic archaeon Pyrobaculum calidifontis. / Wakamatsu, Taisuke; Higashi, Chisato; Ohmori, Taketo; Doi, Katsumi; Ohshima, Toshihisa.

In: Extremophiles, Vol. 17, No. 3, 19.03.2013, p. 379-389.

Research output: Contribution to journalArticle

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