PURIFICATION AND CHARACTERIZATION OF PEA SEEDLING AMINE OXIDASE FOR CRYSTALLIZATION STUDIES

Pea (Pisum sativum L.) seedling amine oxidase (EC 1.4.3.6) is the firs t amine oxidase to be crystallized that diffracts to atomic resolution (2.5 Angstrom). Extensive modifications of a published purification p rocedure were necessary to obtain protein that would give diffraction- quality crystals. Here we report the improved purification and also us e this high-purity protein to reexamine some fundamental characteristi cs of pea seedling amine oxidase. The extinction coefficient at 280 nm (epsilon(280)(1%)) and the molecular mass of the protein are investig ated by a variety of techniques, yielding epsilon(280)(1%) = 20 cm(-1) and a mass of 150 +/- 6 kD. In addition, the stoichiometry of the met al and organic cofactors, Cu(II) and 6-hydroxy dopa (Topa) quinone, re spectively, is examined. The ratio of Cu(II):Topa:protein monomer is f ound to be 1:1:1.