L-Phenylalanine ammonia-lyase (EC 4.3.1.5) was purified from radish cotyledons. The adsorption of the enzyme on L-phenylalanine-Sepharose 4B was nonspecific; the present data disprove the conclusions reached previously (Blondel, J.D., Huault, C., Faye, L., Rollin, P. and Cohen, P. (1973) FEBS Lett. 36, 239-244). 2. The apparent molecular weight of the enzyme fluctuated according to pH and ionic strength. A mean value of 290 000 was determined for the 'native' form. The activation energy was 54.4 kJ/mol. L- and D-phenylalanine protected the lyase from sodium borohydride denaturation. 3. Two Michaelis constants, KHm = 1.5 x 10(-5) M and KLm = 9.5 x 10(-5) M, were determined. The Hill coefficient value (h) was 0.48. This coefficient was 0.6 for cinnamate inhibition. These results suggest that phenylalanine ammonia-lyase in radish cotyledons is regulated by a negative cooperativity mechanism. 4. The phytochrome-mediated increase in phenylalanine ammonia-lyase activity was investigated by density labelling with 2H2O followed by isopycnic centrifugation in KBr gradients. An apparent half-life of 3.5 h was found for both the enzyme extracted from etiolated or far-red irradiated cotyledons. 5. Provided that phytochrome does not control the availability of labelled amino acids for protein synthesis, the present results are consistent with a far-red light-induced increase in the rate of synthesis of the lyase in radish cotyledons.
