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Identification and Characterization of an Efficient Phenylalanine Ammonia-Lyase from Photorhabdus luminescens

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Abstract

A putative aromatic amino acid ammonia-lyase gene (named Pl-pal) was discovered in Photorhabdus luminescens DSM 3368. BLAST and phylogenetic analyses predicted that this enzyme is a histidine ammonia-lyase, whereas sequence alignment suggested that it is more likely a phenylalanine ammonia-lyase (PAL). This gene was amplified from P. luminescens and expressed in Escherichia coli BL21(DE3). The function of Pl-PAL (58 kDa) was characterized by in vitro enzymatic reactions with l-phenylalanine (l-Phe), l-tyrosine (l-Tyr), l-histidine (l-His), and l-tryptophan (l-Trp). Pl-PAL can convert l-Phe and l-Tyr to trans-cinnamic acid and p-coumaric acid, respectively, but had no function on l-His and l-Trp. The optimum temperature and pH were determined to be 40 °C and 11.0, respectively. Under the optimal conditions, Pl-PAL had a kcat/Km value of 0.52 s−1 mM−1 with l-Phe as the substrate, while only 0.013 s−1 mM−1 for l-Tyr. Therefore, the primary function of Pl-PAL was determined to be PAL. The Pl-pal-harboring E. coli strain was used as a whole-cell biocatalyst to produce trans-cinnamic acid from l-Phe. The overall molar conversion rate and productivity were 65.98% and 228.10 mg L−1 h−1, respectively, after the cells were repeatedly utilized 7 times. This work thus provides a promising strain for industrial production of trans-cinnamic acid.

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Data availability

All relevant data are included in this paper. The nucleotide and protein sequences can be accessed in NCBI GenBank based on the accession numbers. Materials are available upon reasonable request to the corresponding author.

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  1. Department of Biological Engineering, Utah State University, 4105 Old Main Hill, Logan, UT, 84322, USA

    Fang Zhang, Jie Ren & Jixun Zhan

  2. College of Life Science and Chemistry, Faculty of Environment and Life, Beijing University of Technology, 100 Pingleyuan, Chaoyang District, Beijing, 100124, China

    Fang Zhang

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JZ designed and coordinated the study. FZ and RJ performed the experiments. FZ, RJ, and JZ analyzed the results. FZ and JZ wrote the manuscript. All of the authors read and approved the final manuscript.

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Correspondence to Jixun Zhan.

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Zhang, F., Ren, J. & Zhan, J. Identification and Characterization of an Efficient Phenylalanine Ammonia-Lyase from Photorhabdus luminescens. Appl Biochem Biotechnol 193, 1099–1115 (2021). https://doi.org/10.1007/s12010-020-03477-6

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