Characterization of 1-aminocyclopropane-1-carboxylate oxidase partially purified from cherimoya fruit

Abstract

The enzyme 1-aminocyclopropane-1-carboxylate (ACC) oxidase was extracted in a soluble form from mesocarp of ripe cherimoya fruit. ACC oxidase purification to near homogeneity was carried out in three chromatographic steps: anion exchange, chromatofocusing, and gel filtration. The molecular mass of the purified enzyme was estimated to be 66 kDa by gel filtration, 62 kDa by native PAGE, and 35 kDa by SDS-PAGE, indicating that the enzyme could be active as a dimer. An isoelectric point at pH 4.35 was estimated by chromatofocusing. The activity of semipurified enzyme eluted from Mono Q columns required Fe2+, sodium bicarbonate, and ascorbate (Km = 6.5 mM). The pH optimum was at 7.4 and the apparent Km with respect to ACC was 82 μM in the absence of added sodium bicarbonate (194 μM in the presence bicarbonate). The particular characteristics of the purified enzyme in relation to the primitive phylogenetic position of cherimoya are discussed.