Purification and characterization of aromatic amine dehydrogenase from Alcaligenes xylosoxidans.
Aromatic amine dehydrogenase was purified and characterized from Alcaligenes xylosoxidans IFO13495 grown on beta-phenylethylamine. The molecular mass of the enzyme was 95.5 kDa. The enzyme consisted of heterotetrameric subunits (alpha2beta2) with two different molecular masses of 42.3 kDa and 15.2 kDa. The N-terminal amino acid sequences of the alpha-subunit (42.3-kDa subunit) and the beta-subunit (15.2-kDa subunit) were DLPIEELXGGTRLPP and APAAGNKXPQMDDTA respectively. The enzyme had a quinone cofactor in the beta-subunit and showed a typical absorption spectrum of tryptophan tryptophylquinone-containing quinoprotein showing maxima at 435 nm in the oxidized form and 330 nm in the reduced form. The pH optima of the enzyme activity for histamine, tyramine, and beta-phenylethylamine were the same at 8.0. The enzyme retained full activity after incubation at 70 degrees C for 40 min. It readily oxidized various aromatic amines as well as some aliphatic amines. The Michaelis constants for phenazine methosulfate, beta-phenylethylamine, tyramine, and histamine were 48.1, 1.8, 6.9, and 171 microM respectively. The enzyme activity was strongly inhibited by carbonyl reagents. The enzyme could be stored without appreciable loss of enzyme activity at 4 degrees C for one month at least in phosphate buffer (pH 7.0).
Kondo
Tetsuya
T
Food Research Center, Aichi Industrial Technology Institute, Aichi Prefectural Government, Nishi-ku, Nagoya, Japan. PXF00674@nifty.ne.jp
Kondo
Emi
E
Maki
Hitomi
H
Yasumoto
Kyoden
K
Takagi
Kazuyoshi
K
Kano
Kenji
K
Ikeda
Tokuji
T
D000421:Alcaligenes / Q000201:enzymology*; D000588:Amines / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D001483:Base Sequence; D016227:Benzoquinones; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010084:Oxidation-Reduction; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D021122:Protein Subunits
D000588:Amines; D001426:Bacterial Proteins; D016227:Benzoquinones; D021122:Protein Subunits; C004532:quinone; D000587:Oxidoreductases Acting on CH-NH Group Donors; C037193:aromatic amine dehydrogenase