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N ‐Methyl‐ l ‐amino acid dehydrogenase from Pseudomonas putida
Author(s) -
Mihara Hisaaki,
Muramatsu Hisashi,
Kakutani Ryo,
Yasuda Mari,
Ueda Makoto,
Kurihara Tatsuo,
Esaki Nobuyoshi
Publication year - 2005
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/j.1742-4658.2004.04541.x
Subject(s) - pseudomonas putida , nad+ kinase , biochemistry , dehydrogenase , formate dehydrogenase , amino acid , oxidoreductase , enzyme , chemistry , cofactor , escherichia coli , stereochemistry , biology , gene
We found N ‐methyl‐ l ‐amino acid dehydrogenase activity in various bacterial strains, such as Pseudomonas putida and Bacillus alvei , and cloned the gene from P. putida ATCC12633 into Escherichia coli . The enzyme purified to homogeneity from recombinant E. coli catalyzed the NADPH‐dependent formation of N‐alkyl‐ l ‐amino acids from the corresponding α‐oxo acids (e.g. pyruvate, phenylpyruvate, and hydroxypyruvate) and alkylamines (e.g. methylamine, ethylamine, and propylamine). Ammonia was inert as a substrate, and the enzyme was clearly distinct from conventional NAD(P)‐dependent amino acid dehydrogenases, such as alanine dehydrogenase (EC 1.4.1.1). NADPH was more than 300 times more efficient than NADH as a hydrogen donor in the enzymatic reductive amination. Primary structure analysis revealed that the enzyme belongs to a new NAD(P)‐dependent oxidoreductase superfamily, the members of which show no sequence homology to conventional NAD(P)‐dependent amino acid dehydrogenases and opine dehydrogenases.