Expression and properties of the highly alkalophilic phenylalanine ammonia-lyase of thermophilic Rubrobacter xylanophilus

Klaudia Kovaćs, Gergely Bańóczi, Andrea Varga, Izabella Szabó, András Holczinger, Gábor Hornyánszky, Imre Zagyva, Csaba Paizs, Beáta G. V́ertessy, Lászĺo Poppe

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19 Citations (Scopus)


The sequence of a phenylalanine ammonia-lyase (PAL; EC: of the thermophilic and radiotolerant bacterium Rubrobacter xylanophilus (RxPAL) was identified by screening the genomes of bacteria for members of the phenylalanine ammonia-lyase family. A synthetic gene encoding the RxPAL protein was cloned and overexpressed in Escherichia coli TOP 10 in a soluble form with an N-terminal His6-tag and the recombinant RxPAL protein was purified by Ni-NTA affinity chromatography. The activity assay of RxPAL with L-phenylalanine at various pH values exhibited a local maximum at pH 8.5 and a global maximum at pH 11.5. Circular dichroism (CD) studies showed that RxPAL is associated with an extensive α-helical character (far UV CD) and two distinctive near-UV CD peaks. These structural characteristics were well preserved up to pH 11.0. The extremely high pH optimum of RxPAL can be rationalized by a three-dimensional homology model indicating possible disulfide bridges, extensive salt-bridge formation and an excess of negative electrostatic potential on the surface. Due to these properties, RxPAL may be a candidate as biocatalyst in synthetic biotransformations leading to unnatural L- or D-amino acids or as therapeutic enzyme in treatment of phenylketonuria or leukemia.

Original languageEnglish
Article numbere85943
JournalPloS one
Issue number1
Publication statusPublished - Jan 27 2014

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

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