The di- and poly-amine oxidases of higher plants.

Diamine oxidase Enzymes which oxidize diamines occur sporadically throughout the plant kingdom (Smith, 1980, 1985a), though they are particularly active in the Leguminosae. The DAO found in pea seedlings (Pisum satiuum) was the subject of early investigations by Werle in Germany (Werle et al., 1961) and by Mann and his co-workers in England (Hill & Mann, 1968). This DAO was purified to homogeneity and shown to contain copper which was readily removed from the enzyme by dialysis against chelating agents, with consequent inactivation. Activity could be restored on addition of small quantities of Cu2+. This property was exploited in the development of an extremely sensitive and highly specific method for the estimation of copper (Hill, 1973). All plant DAOs studied so far are dimers. Several have been shown to contain two copper atoms and one carbonyl residue per mol of enzyme (Nylin & Szybek, 1974; Kluetz et al., 1 9 8 0 ~ ; Matsuda & Suzuki, 1981 ; Yanagisawa et al., 1981 ; Floris et al., 1 9 8 3 ~ ; Rinaldi et al., 1983) though attempts to demonstrate the presence of pyridoxal phosphate have so far failed, and the nature of the carbonyl residue is still unresolved. Recent work on the coppercontaining enzyme, bovine serum amine oxidase, has shown the presence of pyrroloquinoline quinone (Lobenstein-Verbeek et al., 1984). It remains to be seen if this is also the carbonyl cofactor for the pea seedling DAO. Estimates of the M , of the dimeric form of the pea seedling DAO are in the range 170000-185000 (McGowan & Muir, 1971 ; Macholan & Haubrovi, 1976; Kluetz et al., 1 9 8 0 ~ ; Yanagisawa et al., 1981). Pea seedlings are the most active source of DAO, exceeding the classical hog kidney by 105 times in terms of crude material, and by 58 times in terms of purified enzyme (Smith, 1979). Moreover, the pea seedling enzyme is very stable during long-term storage and in assay. For these reasons the pea seedling DAO may be utilized for estimating amines. In one example of this, an arginine decarboxylase assay was developed in which the product of the decarboxylase (i.e. agmatine) was oxidized in the assay incubate by pea seedling DAO, and the peroxide produced was then estimated by means of the peroxidative oxidation of guaiacol (Smith, 1979). This assay is considerably more rapid and convenient than the conventional isotopic method more usually adopted for amino acid decarboxylase assay. The sensitivity could be greatly increased by utilizing peroxidase substrates which fluoresce on oxidation (Smith, 1983~). A method has also been developed for estimating lysine and arginine, by using membrane-bound DAO and the respective amino acid decarboxylases, in association with an oxygen electrode (Macholan, 1978). A mathematical model has been established for amine estimation by this electrode system (Toul & Macholan, 1975). When investigating amine metabolism in plants, especially in the Leguminosae, it is important to be aware of the presence of DAO, since this may cause artefactual reactions catalysed by peroxidase (Suresh & Adiga, 1977). However, in viuo the peroxide formed may be utilized by the plant to affect natural metabolic pathways such as uric acid degradation (Tajima ef al., 1983). Within the Leguminosae, DAOs with properties similar to those of the pea seedling enzyme have beeen found in