Molecular characterization of a phenylalanine ammonia-lyase gene (BoPAL1) from Bambusa oldhamii

• Published in: Molecular biology reports Vol. 38; no. 1; pp. 283 - 290
• Main Authors: Hsieh, Lu-Sheng, Hsieh, Yi-Lin, Yeh, Chuan-Shan, Cheng, Chieh-Yang, Yang, Chien-Chih, Lee, Ping-Du
• Format: Journal Article
• Language: English
• Published: Dordrecht Dordrecht : Springer Netherlands 2011; Springer Netherlands; Springer Nature B.V
• Subjects: 5' Flanking Region - genetics; Ammonia; Angiosperms; Animal Anatomy; Animal Biochemistry; Bamboo (Bambusa oldhamii); Bambusa; Bambusa - enzymology; Bambusa - genetics; Base Sequence; Biomedical and Life Sciences; Chromatography; Chromatography, Affinity; Cloning; Cloning, Molecular; double prime P elements; Enzymes; Filtration; Flowers & plants; Gene expression; Genes, Plant - genetics; Histology; Kinetics; Life Sciences; Models, Molecular; Molecular biology; molecular cloning; Molecular Sequence Data; Morphology; Open reading frames; pal gene; pH effects; Phenylalanine ammonia-lyase; Phenylalanine ammonia-lyase (PAL); Phenylalanine Ammonia-Lyase - chemistry; Phenylalanine Ammonia-Lyase - genetics; phenylpropanoids; Pichia - metabolism; Pichia pastoris; Plant Proteins - chemistry; Plant Proteins - genetics; Recombinant Proteins - isolation & purification; Regulatory Sequences, Nucleic Acid - genetics; Species Specificity; TAIL-PCR; Temperature effects; ); Molecular cloning; TAIL-PCR; Gene expression; Phenylalanine ammonia-lyase (PAL); Bamboo
• ISSN: 0301-4851; 1573-4978
• DOI: 10.1007/s11033-010-0106-2

Phenylalanine ammonia-lyase is the first enzyme of general phenylpropanoid pathway. A PAL gene, designated as BoPAL1, was cloned from a Bambusa oldhamii cDNA library. The open reading frame of BoPAL1 was 2,139 bp in size and predicted to encode a 712-amino acid polypeptide. BoPAL1 was the first intronless PAL gene found in angiosperm plant. Several putative cis-acting elements such as P box, GT-1motif, and SOLIPs involved in light responsiveness were found in the 5′-flanking sequence of BoPAL1 which was obtained by TAIL-PCR method. Recombinant BoPAL1 protein expressed in Pichia pastoris was active. The optimum temperature and pH for BoPAL1 activity was 50°C and 9.0, respectively. The molecular mass of recombinant BoPAL1 was estimated as 323 kDa using gel filtration chromatography and the molecular mass of full-length BoPAL was about 80 kDa, indicating that BoPAL1 presents as a homotetramer. The K m and k cat values of BoPAL1 for L-Phe were 1.01 mM and 10.11 s⁻¹, respectively. The recombinant protein had similar biochemical properties with PALs reported in other plants.