Partial purification and properties of phenylalanine ammonia-lyase from leaves of mango.
Ananthanarayanan T. V., Chacko E. K.
Author Affiliation: Indian Institute of Horticultural Research, Bangalore 560?080, India.
: 483-490
Abstract : Finely divided Alphonso mango leaves were extracted with cooled acetone:n-butanol (9:1). The residual powder was blended with 0.05 M borate buffer (pH 8.8) and hydrated polyvinylpyrolidone, and then hydrolysed, filtered, and centrifuged to give the crude enzyme extract. The extract was further purified by dialysis, fractionation by precipitation with ammonium sulphate (40%, 40-60% and 60-80%), and DEAE cellulose ion-exchange chromatography. Total protein content, L-phenylalanine ammonia-lyase (PAL) activity (determined by the magnitude of absorption at 290 nm, at pH 8.8) and tyrosine ammonia-lyase (TAL) activity (absorption of p-coumarate, the product of deamination of L-tyrosine, at 308 nm, at pH 8.8) were determined on the different fractions. The ratio of PAL to TAL activity was the same (2.9-3.0) for all fractions. PAL was activated by sulphydryl compounds and inhibited by ZnCl2 (65% at 10-3 M) but not by MgCl2 (2 × 10-4 M) or CaCl2 (3 × 10-3 M). PAL activity was higher in newer purple-brown leaves than in older green leaves. PAL is active up to 58°C, with a maximum at pH 8.8, and TAL is active up to 46°, with a maximum at pH 7.5.