Purification, cloning, and three-dimensional structure prediction of Micrococcus luteus FAD-containing tyramine oxidase.
The FAD-containing tyramine oxidase enzyme and gene from the Gram (+) bacterium Micrococcus luteus were isolated, and computer prediction was used to propose a preliminary 3D model of the protein. A 2.8-kb Sau3AI fragment containing the structural gene of tyramine oxidase was cloned from a M. luteus genomic DNA library. The 1332 bp gene encodes a protein of 443 amino acids, with a calculated molecular mass of 49.1 kDa. The enzyme was found to be a homodimer with a molecular weight of 49,000. It oxidizes tyramine, adrenaline, 3-hydroxytyramine, dopamine, and noradrenaline, and was reversibly inhibited by FAD-containing monoamine oxidase A and B specific inhibitors. Sequence comparison show that tyramine oxidase is smaller than other FAD-amine oxidases but that it contains well-conserved amino acid residues reported in all other FAD-amine oxidases. A hypothetical three-dimensional structure of tyramine oxidase has also been proposed based on secondary structure predictions, threading, and comparative modeling.
Roh
J H
JH
Research Institute of Innovative Technology for the Earth, Soraku-gun, Kyoto, Japan.
Wouters
J
J
Depiereux
E
E
Yukawa
H
H
Inui
M
M
Minami
H
H
Suzuki
H
H
Kumagai
H
H
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000032:analysis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry; D016982:Micrococcus luteus / Q000737:chemistry / Q000235:genetics*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008995:Monoamine Oxidase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D011487:Protein Conformation; D013379:Substrate Specificity; D014158:Transcription, Genetic
D004269:DNA, Bacterial; D004791:Enzyme Inhibitors; D005182:Flavin-Adenine Dinucleotide; D008995:Monoamine Oxidase