The fungus Gibberella fujikuroi produces copper/topaquinone-containing amine oxidase when induced by N-butylamine.
Crude extract of Gibberella fujikuroi AKU 3802 mycelium induced with n-butylamine showed a single amine oxidase activity band in a non-denaturing gel that cross-reacted with the antibody against copper/topaquinone-containing amine oxidase from Aspergillus niger. The enzyme was purified by a procedure involving four chromatographic steps. Purified enzyme was pink with an absorption maximum at 490 nm. Molecular mass of 135 kDa estimated by gel chromatography and 70 kDa found by SDS-PAGE confirmed the dimeric structure of the enzyme. The enzyme readily oxidized n-hexylamine, n-butylamine, benzylamine and histamine, but not spermine or spermidine. Inactivation by carbonyl reagents and copper chelators suggested the presence of a copper/topaquinone cofactor. Spectrophotometric titration by p-nitrophenylhydrazine showed one reactive carbonyl group per subunit and redox-cyclic quinone staining confirmed the presence of a quinone cofactor. pH-dependent shift of the absorption spectrum of the enzyme p-nitrophenylhydrazone (465 nm at neutral to 580 nm at alkaline pH) supports the identity of the cofactor, with topaquinone. The N-terminal amino acid sequence of the enzyme showed high similarity to other microbial copper/topaquinone-containing amine oxidases.
Frébort
I
I
Department of Biological Chemistry, Faculty of Agriculture, Yamaguchi University, Japan.
Matsushita
K
K
Adachi
O
O
D006631:Amine Oxidase (Copper-Containing) / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D002082:Butylamines; D004295:Dihydroxyphenylalanine / Q000031:analogs & derivatives / Q000032:analysis; D005874:Gibberella / Q000201:enzymology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016415:Sequence Alignment; D013379:Substrate Specificity
D002082:Butylamines; D004295:Dihydroxyphenylalanine; C067626:6-hydroxydopa quinone; D006631:Amine Oxidase (Copper-Containing); C032462:n-butylamine