title: Structural insights into the substrate specificity of bacterial copper amine oxidase obtained by using irreversible inhibitors.
abstract: Copper amine oxidases (CAOs) catalyse the oxidation of various aliphatic amines to the corresponding aldehydes, ammonia and hydrogen peroxide. Although CAOs from various organisms share a highly conserved active-site structure including a protein-derived cofactor, topa quinone (TPQ), their substrate specificities differ considerably. To obtain structural insights into the substrate specificity of a CAO from Arthrobacter globiformis (AGAO), we have determined the X-ray crystal structures of AGAO complexed with irreversible inhibitors that form covalent adducts with TPQ. Three hydrazine derivatives, benzylhydrazine (BHZ), 4-hydroxybenzylhydrazine (4-OH-BHZ) and phenylhydrazine (PHZ) formed predominantly a hydrazone adduct, which is structurally analogous to the substrate Schiff base of TPQ formed during the catalytic reaction. With BHZ and 4-OH-BHZ, but not with PHZ, the inhibitor aromatic ring is bound to a hydrophobic cavity near the active site in a well-defined conformation. Furthermore, the hydrogen atom on the hydrazone nitrogen is located closer to the catalytic base in the BHZ and 4-OH-BHZ adducts than in the PHZ adduct. These results correlate well with the reactivity of 2-phenylethylamine and tyramine as preferred substrates for AGAO and also explain why benzylamine is a poor substrate with markedly decreased rate constants for the steps of proton abstraction and the following hydrolysis.
authors: Department of Biochemistry, Osaka Medical College, Osaka 569-8686, Japan.
authors: Takeshi
authors: T
authors: Murakawa
authors: Hideyuki
authors: H
authors: Hayashi
authors: Masayasu
authors: M
authors: Taki
authors: Yukio
authors: Y
authors: Yamamoto
authors: Yoshiaki
authors: Y
authors: Kawano
authors: Katsuyuki
authors: K
authors: Tanizawa
authors: Toshihide
authors: T
authors: Okajima
mesh_terms: D006631:Amine Oxidase (Copper-Containing) / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D001173:Arthrobacter / Q000201:enzymology / Q000378:metabolism; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D002384:Catalysis; D004295:Dihydroxyphenylalanine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000378:metabolism; D006834:Hydrazines / Q000737:chemistry / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D010084:Oxidation-Reduction; D012545:Schiff Bases; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D006834:Hydrazines; D012545:Schiff Bases; C029424:hydrazine; D004295:Dihydroxyphenylalanine; C067626:6-hydroxydopa quinone; D006631:Amine Oxidase (Copper-Containing)