Purification and characterization of a copper-containing amine oxidase from Mycobacterium sp. strain JC1 DSM 3803 grown on benzylamine.
A bacterial semicarbazide-sensitive amine oxidase (SSAO) was purified and characterized from Mycobacterium sp. strain JC1 DSM 3803 grown on benzylamine. During the purification procedures, the enzyme was tending to aggregate and exhibited heterogeneity in native PAGE. The heterogeneous forms having amine oxidase (AO) activity could be separated by their native molecular weights using gel-filtration chromatography. Most of the AOs behaved as dimers (M(r) 150,000) composed of a 75-kDa subunit, but some aggregated to form tetramers (M(r) 300,000). Besides their native molecular weight, subunit composition and V(max) value, both forms (dimer and tetramer) have almost identical biochemical properties (e.g. subunit size, optimum pH and temperature, activation energy, K(m) value on benzylamine, substrate and inhibitor specificities). When AO activity was observed by activity staining, the best-oxidized substrate was benzylamine, although the AO also oxidized tyramine and histamine. The AO was strongly inhibited by semicarbazide and isoniazid, but KCN did not affect its activity. The purified enzyme was shown to contain 2.39 mol of copper per mole of subunit, but there were no evidences of topaquinone co-factor involvement, when tested by absorption spectrum analysis and redox-cycling staining for quinoprotein detection.
Lee
Hyun-Il
HI
Laboratory of Biochemistry, Graduate School of Medicine, Konkuk University, Chungju 380-701, Korea.
Kim
Young Min
YM
Ro
Young Tae
YT
D006631:Amine Oxidase (Copper-Containing) / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D001596:Benzylamines / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008970:Molecular Weight; D009161:Mycobacterium / Q000201:enzymology* / Q000254:growth & development; D021122:Protein Subunits / Q000737:chemistry; D013379:Substrate Specificity
D001426:Bacterial Proteins; D001596:Benzylamines; D004791:Enzyme Inhibitors; D021122:Protein Subunits; C030796:benzylamine; D006631:Amine Oxidase (Copper-Containing)