Purification and spectral characterization of a paraperoxidase from Cucurbita pepo ripe fruits

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The isolation and purification of the two major anionic (A 1) and cationic (C 1) peroxidase isoenzymes extracted from the ripe fruit of Curcubita pepo have been described. The anionic isoenzyme A 1 has spectral properties very similar to those of normal horseradish peroxidase isoenzymes. The cationic isoenzyme C 1 has a paraperoxidase nature and has been characterized by electronic and circular dichroism spectroscopy, together with several of its derivatives. The spectra suggest that native isoenzyme C 1 may exist as equilibrium mixture of high-spin and low-spin forms, but such equilibria are apparently absent in the various derivatives of the enzyme. Careful comparison of the spectra of the native isoenzyme C 1 and its reduced form with those of the corresponding cyanide derivatives seems to exclude the possibility that the paraperoxidase character could arise from a cyanide-bound form of the enzyme. The spectra of isoenzyme C 1 recorded in the presence of various substrates and H 2O 2 indicate an accumulation of compound III.