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PURIFICATION AND CHARACTERIZATION OF THREE POLYPHENOL OXIDASE ISOZYMES FROM LOBSTER ( HOMARUS AMERICANUS )
Author(s) -
OPOKUGYAMFUA A.,
SIMPSON B.K.
Publication year - 1992
Publication title -
journal of food biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.507
H-Index - 47
eISSN - 1745-4514
pISSN - 0145-8884
DOI - 10.1111/j.1745-4514.1992.tb00453.x
Subject(s) - polyphenol oxidase , isozyme , isoelectric point , homarus , chemistry , substrate (aquarium) , ion chromatography , biochemistry , enzyme , isoelectric focusing , catechol oxidase , chromatography , biology , peroxidase , ecology , crustacean
Three isozymes of polyphenol oxidase (PPO I, PPO II and PPO III) were purified from lobster (Homarus americanus) by ion‐exchange chromatography and preparative isoelectric focusing using a Rotofor cell. The purified isozymes migrated as single protein bands in polyacrylamide gels with R f values corresponding to molecular weights of 32,180, 35,480 and 39,300, respectively. The pI values of the PPO isozymes were 3.89, 4.26 and 4.54, respectively. PPO I was most active at pH 6.5 and most stable from pH 6.0 to 7.0; PPO II was most active within the pH range 6.0 to 7.0, and most stable within the pH range 4.0 to 9.0; while PPO III was most active at pH 7.0 and most stable in the pH range 6.0 to 8.0. The temperature optimum for the PPO‐dihydroxyphenylalanine oxidation reaction was 35C with PPO I and 45C with PPO II or III. Lobster PPO I lost about 30% of its initial activity after 30 min incubation at 45C, while PPO II and II retained virtually all their activity after the same heat treatment. The catalytic specificities of the PPO isozymes were relatively higher with dihydroxyphenylalanine as substrate than with chlorogenic acid.