Purification and Characterization of Phenylalanine Ammonia Lyase from Trichosporon cutaneum

Abstract EN

Trichosporon cutaneum phenylalanine ammonia lyase was selected as a model to investigate the dual substrate activity of this family of enzymes.

Sequencing of the PAL gene identified an extensive intron region at the N-terminus.

Five amino acid residues differing from a prior report were identified.

Highest Phe : Tyr activities (1.6 ±0.3 : 0.4±0.1 μmol/h g wet weight) were induced by Tyr.

The enzyme has a temperature optimum of 32°C and a pH optimum of 8–8.5 and shows no metal cofactor dependence.

Michaelis-Menten kinetics (Phe, Km 5.0 ± 1.1 mM) and positive allostery (Tyr, K′ 2.4 ± 0.6 mM, Hill coefficient 1.9±0.5) were observed.

Anion exchange chromatography gave a purification fold of 50 with 20% yield.

The His-Gln motif (substrate selectivity switch region) indicates the enzyme’s ability to act on both substrates.