A novel high activity cationic ascorbate peroxidase from tea (Camellia sinensis) - A class III peroxidase with unusual substrate specificity

A cationic class III peroxidase (TcAPX II) with the highest reported specif ic activity (k(cat) = 1,500 mu mol min(-1) mg(-1)) for ascorbate as reducin g substrate has been isolated from freshly picked tea leaves (Camellia sine nsis) in 45 % glycosylated, 55 % non-glycosylated forms. TcAPX II exhibits important structure-function differences with respect to not only conventio nal class I (e.g. pea cytosolic ascorbate peroxidase) and class III peroxid ases (e.g. horseradish peroxidase) but also to another recently characteris ed class III ascorbate specific enzyme, TcAPX I [Kvaratskhelia et al. Plant Physiol. 144, 1237-1245 (1997)]. TcAPX II has a high preference for ascorb ate as a reducing substrate, while TcAPX I oxidises ascorbate and organic p henols at 10-fold lower, but comparable rates. Hydrogen peroxide (100-4,000 fold excess) reacts with the ferric and compound II states of TcAPX II to yield compound II and an inactive type P670 species with no detectable comp ound III formation. The inactivation rate is comparable with that of horser adish peroxidase but significantly lower than that of pea cytosolic APX. Th ese data together with the instability of TcAPX II compound I (t(1/2) = 5 s ec) in the absence of added reducing substrate, suggest that TcAPX II is pr otected from H2O2 induced inactivation by a catalase like reaction. Partial sequence data for TcAPX II show that recognised structural similarities be tween class I ascorbate peroxidases and yeast cytochrome c peroxidase (the archetypal class I peroxidase) are not essential for ascorbate peroxidase a ctivity. TcAPX II is a distinct class III peroxidase of generic interest be cause of its potential to act as a key antioxidant in aerobic stress respon se in planta.