Characterization of polyphenol oxidase from aerial roots of an orchid, Aranda `Christine 130ʹ

Four isoforms of polyphenol oxidase (PPO) were demonstrated in the aerial roots of a tropical orchid, Aranda `Christine 130ʹ. They were extracted at neutral pH and purified to homogeneity as judged by SDS-gel electrophoresis. Purification was achieved by a combination of Triton X-114 treatment, temperature phase partitioning, gel filtration chromatography, ion-exchange separation and chromatofocusing. Two of the isoforms, designated PPOa and PPOd, differed in their N-terminal sequence, tryptic peptide map and substrate affinity for (+)-catechin, but exhibited similarity in their molecular mass under denaturing conditions, pH optimum and kinetic behaviour toward 4-methyl catechol. The other two isoforms, PPOb and PPOc, were identical to PPOa and PPOd, respectively, in terms of their N-terminal sequence, substrate preference and pH maximum, but were different with regard to their molecular mass under denaturing conditions. These four isoforms differed in their isoelectric point.