L-ORNITHINE DECARBOXYLASE FROM HAFNIA-ALVEI HAS A NOVEL L-ORNITHINE OXIDASE ACTIVITY

A novel activity producing gamma-aminobutyric acid (GABA) from L-ornit hine in the presence of NAD(P)(+) was found in the crude extract of L- ornithine-induced Hafnia alvei, in addition to L-ornithine decarboxyla se (ODC) activity. The reaction system for the former activity consist ed of two enzymes, L-ornithine oxidase (decarboxylating, GOD) and gamm a-aminobutyraldehyde (GABL) dehydrogenase (GDH). OOD catalyzed the con version of L-ornithine into GABL, CO2, NH3, and H2O2 in the presence o f O-2, and GDH dehydrogenated GABL to GABA in the presence of NAD(P)(). OOD, purified to homogeneity, had a high ODC activity and the activ ity ratio of ODC to OOD was almost constant throughout the purificatio n (ODC/OOD = 160 : 1). The molecular mass of the OOD was about 230 kDa , probably consisting of three identical subunits of a 77 kDa peptide, and OOD had an absorption maximum at 420 nm as well as at 278 nm, the specific absorption for an enzyme containing pyridoxal phosphate (PLP ). The content of PLP was estimated at about 1 mol per subunit. OOD wa s specific to L-ornithine, and other L-amino acids and polyamines incl uding putrescine were inert. The enzyme was activated by PLP, but not by pyridoxamine 5'-phosphate, FAD, FMN, or pyrroloquinoline quinone, a nd it was inactivated by hydrazine, semicarbazide, and hydroxylamine. The holoenzyme can be resolved to the apoenzyme by incubation with hyd roxylamine, and reconstituted with PLP. These properties of OOD were a lmost the same as those of ODC separately purified to homogeneity from H. alvei. Zn2+ and Cu2+, butane-dione, and sodium borohydride inhibit ed both OOD and ODC in a similar manner. The OOD reaction required O-2 and only the ODC reaction proceeded under anaerobic conditions. The s ubstitution of air for oxygen in the reaction vessel and the addition of catalase-H,Oz enhanced only the OOD reaction, resulting in an incre ase of the ratio of OOD/ODC to 1:30 and 1:4.1, respectively. These res ults suggested that OOD and ODC are identical and that the former is a side reaction of the latter in the presence of O-2.