PURIFICATION, PROPERTIES AND PARTIAL AMINO-ACID-SEQUENCE OF 1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID OXIDASE FROM APPLE FRUITS

The enzyme which converts 1-aminocyclopropane-1-carboxylic acid (ACC) into ethylene, ACC oxidase, has been isolated from apple fruits (Malus x domestica Borkh. cv. Golden Delicious), and for the first time stab ilized in vitro by 1,10-phenanthroline and purified 170-fold to homoge neity in a five-step procedure. The sodium dodecyl sulfate-denatured a nd native proteins have similar molecular weights (approx. 40 kDa) ind icating that the enzyme is active in its monomeric form. Antibodies ra ised against a recombinant ACC oxidase overproduced in Escherichia col i from a tomato cDNA recognise the apple-fruit enzyme with high specif icity in both crude extracts and purified form. Glycosylation appears to be absent because of (i) the lack of reactivity towards a mixture o f seven different biotinylated lectins and (ii) the absence of N-linke d substitution at a potential glycosylation site, in a sequenced pepti de. Phenylhydrazine and 2-methyl-1-2-dipyridyl propane do not inhibit activity, indicating that ACC oxidase is not a prosthetic-heme iron pr otein. The partial amino-acid sequence of the native protein has stron g homology to the predicted protein of a tomato fruit cDNA demonstrate d to encode ACC oxidase.