Gene cloning and structure characterization of phenylalanine ammonia-lyase from Arisaema heterophyllum

Phenylalanine ammonia-lyase (PAL) is the first key enzyme in the synthesis of flavonoids and the rate-limiting enzyme of the phenylpropanoid pathway, which is associated with secondary metabolites, growth and development, as well as defense of plants. In this study, the mRNA of Arisaema heterophyllum Blume was used as a template and the full-length open reading frame (ORF) of the phenylalanine ammonia-lyase gene (AhPAL) was amplified by real-time polymerase chain reaction (RT-PCR). The structure and physicochemical properties of AhPAL were then analyzed by bioinformatics. Results showed that the ORF of AhPAL was 2,184 bp in length, encoding a protein with 727 amino acids. Furthermore, AhPAL had the closest relationship with PAL of Tulipa fosteriana W. Irving, with a sequence identity of 88%. Structure analysis indicated that AhPAL was a homotypic tetramer, and each monomer consisted of three domains. Among these domains, the MIO domain was highly important for the activity of AhPAL, containing a conserved sequence of the PAL family and Ala-Ser-Gly triad residues. The expression levels of three AhPAL unigenes in the roots, tubers, and leaves were detected by quantitative RT-PCR (qRT-PCR), with higher expression found for all three in the roots compared with that in the leaves and tubers.