Induction of N-hydroxycinnamoyltyramine synthesis and tyramine N-hydroxycinnamoyltransferase (THT) activity by wounding in maize leaves

Both N-p-coumaroyl- and N-feruloyltyramine accumulated in response to wound ing in leaf segments of maize. The amount of N-hydroxycinnamoyltyramines st arted to increase 3-6 h after wounding and peaked at 12 h. Thereafter, the amount of N-p-coumaroyltyramine decreased rapidly, while the N-feruloyltyra mine content remained at a high level. The accumulation of N-hydroxycinnamo yltyramines was accompanied by an increase in the tyramine N-hydroxycinnamo yltransferase (THT) activity. This increase was initially detected 3 h afte r wounding and reached a maximum at 36 h, the level of activity being 40 an d 11 times that in the leaves before wounding and in the control leaves, re spectively. Partial purification of THT from wounded leaves by (NH4)(2)SO4 precipitation and subsequent two steps of anion-exchange chromatography res ulted in a 12.5-fold increase in specific activity. Kinetic studies with th is partially purified enzyme revealed that the best substrates were tyramin e and feruloyl-CoA, although tryptamine and sinapoyl-CoA also efficiently s erved as substrates. The apparent native molecular weight of the enzyme was determined by gel filtration as 40 kDa.