Characterization of a thermostable 2,4-diaminopentanoate dehydrogenase from Fervidobacterium nodosum Rt17-B1.
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2, 4-Diaminopentanoate dehydrogenase (2, 4-DAPDH), which is involved in the oxidative ornithine degradation pathway, catalyzes the NAD(+)- or NADP(+)-dependent oxidative deamination of (2R, 4S)-2, 4-diaminopentanoate (2, 4-DAP) to form 2-amino-4-oxopentanoate. A Fervidobacterium nodosum Rt17-B1 gene, Fnod_1646, which codes for a protein with sequence similarity to 2, 4-DAPDH discovered in metagenomic DNA, was cloned and overexpressed in Escherichia coli, and the gene product was purified and characterized. The purified protein catalyzed the reduction of NAD(+) and NADP(+) in the presence of 2, 4-DAP, indicating that the protein is a 2, 4-DAPDH. The optimal pH and temperature were 9.5 and 85°C, respectively, and the half-denaturation time at 90°C was 38 min. Therefore, the 2, 4-DAPDH from F. nodosum Rt17-B1 is an NAD(P)(+)-dependent thermophilic-alkaline amino acid dehydrogenase. This is the first thermophilic 2, 4-DAPDH reported, and it is expected to be useful for structural and functional analyses of 2, 4-DAPDH and for the enzymatic production of chiral amine compounds. Activity of 2, 4-DAPDH from F. nodosum Rt17-B1 was suppressed by 2, 4-DAP via uncompetitive substrate inhibition. In contrast, the enzyme showed typical Michaelis-Menten kinetics toward 2, 5-diaminohexanoate. The enzyme was uncompetitively inhibited by d-ornithine with an apparent Ki value of 0.1 mM. These results suggest a regulatory role for this enzyme in the oxidative ornithine degradation pathway.
収録刊行物
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- Journal of bioscience and bioengineering
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Journal of bioscience and bioengineering 117 (5), 551-556, 2014-05
Elsevier B.V.
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詳細情報 詳細情報について
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- CRID
- 1050845760721632512
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- NII論文ID
- 110009823129
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- NII書誌ID
- AA11307678
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- ISSN
- 13474421
- 13891723
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- HANDLE
- 2433/187042
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- NDL書誌ID
- 025469334
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- 本文言語コード
- en
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- 資料種別
- journal article
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- データソース種別
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