Coniferyl alcohol oxidase activity of a cell-wall-located class III peroxidase

Coniferyl alcohol oxidase activity was determined in cell walls from hypocotyls of the following species belonging to the family Asteraceae: Calendula officinalis , Callistephus sinensis , Cosmos bipinnanthus , Helianthus annuus , Helianthus debilis and Zinnia elegans . In all the cases studied, coniferyl alcohol oxidase activity was partially located ionically-bound to cell walls and resided in a basic peroxidase, the activity of which was stimulated by H 2 O 2 . This enzymatic activity was insensitive to freezing and was inactivated by high H 2 O 2 concentrations, as tested both in vitro and in situ by using purified cell wall fractions. The peroxidase with coniferyl alcohol oxidase activity was purified from Z. elegans hypocotyls until apparent homogeneity, as checked by SDS-PAGE. It showed a visible spectrum typical of a haem-containing high-spin ferric secretory (class III) plant peroxidase. Coniferyl alcohol oxidase activity of this basic peroxidase constitutes about 0.25% of the activity shown in the presence of H 2 O 2 . The significance of the coniferyl alcohol oxidase activity in vivo was studied in Z. elegans hypocotyls by means of histochemical tests, which revealed that it was located in the H 2 O 2 -producing lignifying xylem cells. The results obtained from the histochemical probes suggest that the coniferyl alcohol oxidase activity of this basic peroxidase is physiologically irrelevant in tissues that accumulate H 2 O 2 , as is the case of the lignifying xylem, where the peroxidase activity of the enzyme favorably competes with the oxidase activity of the enzyme.