A novel phenylalanine ammonia-lyase from Pseudozyma antarctica for stereoselective biotransformations of unnatural amino acids

• Published in: Catalysis today Vol. 366; pp. 185 - 194
• Main Authors: Varga, Andrea, Csuka, Pál, Sonesouphap, Orlavanah, Bánóczi, Gergely, Toşa, Monica Ioana, Katona, Gabriel, Molnár, Zsófia, Bencze, László Csaba, Poppe, László, Paizs, Csaba
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 15.04.2021
• Subjects: Biocatalysis; Phenylalanine ammonia-lyase; Pseudozyma antarctica; Unnatural amino acids; Yeast; Pseudozyma antarctica; Phenylalanine ammonia-lyase; Biocatalysis; Yeast; Unnatural amino acids
• ISSN: 0920-5861; 1873-4308
• DOI: 10.1016/j.cattod.2020.04.002

[Display omitted] •Novel phenylalanine ammonia-lyase for the production of unnatural amino acids.•Superior turnover numbers compared to known phenylalanine ammonia-lyases.•Superior enantioselectivity in the synthesis of unnatural phenylalanines bearing electron withdrawing substituents.•High thermal stability for an enzyme of psychrophilic origin. A novel phenylalanine ammonia-lyase of the psychrophilic yeast Pseudozyma antarctica (PzaPAL) was identified by screening microbial genomes against known PAL sequences. PzaPAL has a significantly different substrate binding pocket with an extended loop (26 aa long) connected to the aromatic ring binding region of the active site as compared to the known PALs from eukaryotes. The general properties of recombinant PzaPAL expressed in E. coli were characterized including kinetic features of this novel PAL with l-phenylalanine (S)-1a and further racemic substituted phenylalanines rac-1b-g,k. In most cases, PzaPAL revealed significantly higher turnover numbers than the PAL from Petroselinum crispum (PcPAL). Finally, the biocatalytic performance of PzaPAL and PcPAL was compared in the kinetic resolutions of racemic phenylalanine derivatives (rac-1a-s) by enzymatic ammonia elimination and also in the enantiotope selective ammonia addition reactions to cinnamic acid derivatives (2a-s). The enantiotope selectivity of PzaPAL with o-, m-, p-fluoro-, o-, p-chloro- and o-, m-bromo-substituted cinnamic acids proved to be higher than that of PcPAL.