title: Phenylalanine ammonia lyase from Arabidopsis thaliana (AtPAL2): A potent MIO-enzyme for the synthesis of non-canonical aromatic alpha-amino acids: Part I: Comparative characterization to the enzymes from Petroselinum crispum (PcPAL1) and Rhodosporidium toruloides (RtPAL).
abstract: Phenylalanine ammonia lyase (PAL) from Arabidopsis thaliana (AtPAL2) was comparatively characterized to the well-studied enzyme from parsley (PcPAL1) and Rhodosporidium toruloides (RtPAL) with respect to kinetic parameters for the deamination and the amination reaction, pH- and temperature optima and the substrate range of the amination reaction. Whereas both plant enzymes are specific for phenylalanine, the bifunctional enzyme from Rhodosporidium toruloides shows KM-values for L-Phe and L-Tyr in the same order of magnitude and, compared to both plant enzymes, a 10-15-fold higher activity. At 30°C all enzymes were sufficiently stable with half-lives of 3.4days (PcPAL1), 4.6days (AtPAL2) and 9.7days (RtPAL/TAL). Very good results for the amination of various trans-cinnamic acid derivatives were obtained using E. coli cells as whole cell biocatalysts in ammonium carbonate buffer. Investigation of the substrate ranges gave interesting results for the newly tested enzymes from A. thaliana and R. toruloides. Only the latter accepts besides 4-hydroxy-CA also 3-methoxy-4-hydroxy-CA as a substrate, which is an interesting intermediate for the formation of pharmaceutically relevant L-Dopa. AtPAL2 is a very good catalyst for the formation of (S)-3-F-Phe, (S)-4-F-Phe and (S)-2-Cl-Phe. Such non-canonical amino acids are valuable building blocks for the formation of various drug molecules.
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, D-52425 Jülich, Germany. Electronic address: alana.dressen@gmx.de.
authors.forename: Alana
authors.initials: A
authors.lastname: Dreßen
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, D-52425 Jülich, Germany. Electronic address: Thomas.Hilberath@uni-duesseldorf.de.
authors.forename: Thomas
authors.initials: T
authors.lastname: Hilberath
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, D-52425 Jülich, Germany. Electronic address: u.mackfeld@fz-juelich.de.
authors.forename: Ursula
authors.initials: U
authors.lastname: Mackfeld
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, D-52425 Jülich, Germany. Electronic address: arne.billmeier@gmx.de.
authors.forename: Arne
authors.initials: A
authors.lastname: Billmeier
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Sciences, Section: Technical Biology, Engler-Bunte-Ring 3, D-76131 Karlsruhe, Germany. Electronic address: jens.rudat@kit.edu.
authors.forename: Jens
authors.initials: J
authors.lastname: Rudat
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, D-52425 Jülich, Germany. Electronic address: ma.pohl@fz-juelich.de.
authors.forename: Martina
authors.initials: M
authors.lastname: Pohl
mesh_terms: D024322:Amino Acids, Aromatic / Q000378:metabolism*; D017360:Arabidopsis / Q000201:enzymology*; D001487:Basidiomycota / Q000201:enzymology*; D005656:Fungal Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007093:Imidazoles; D028528:Petroselinum / Q000201:enzymology*; D010650:Phenylalanine Ammonia-Lyase / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013237:Stereoisomerism
chemical_list: D024322:Amino Acids, Aromatic; D005656:Fungal Proteins; D007093:Imidazoles; D010940:Plant Proteins; D011994:Recombinant Proteins; C447541:methylidene-imidazolone; D010650:Phenylalanine Ammonia-Lyase
keywords: (S)-2-F-Phe (Pubchem CID: 716319); (S)-3-Cl-Phe (Pubchem CID: 85680); (S)-3-F-Phe (Pubchem CID: 9976); (S)-3-OCH(3)-4-OH-Phe (Pubchem CID: 94331; (S)-4-Cl-Phe (Pubchem CID:4652); (S)-4-F-Phe (Pubchem CID: 716312); (S)-Phe (Pubchem CID: 6140); (S)-tyrosine (Pubchem CID 6057); (S)‐2‐Cl-Phe (Pubchem CID85679); Chiral synthesis; MIO enzyme; Phenylalanine ammonia lyase; Substrate range; Tyrosine ammonia lyase; Whole cell biotransformation; trans-cinnamic acid (Pubchem CID: 444539)