Molecular Cloning and Characterization of the Phenylalanine Aminomutase Gene From Taxus baccata L.

Document Type : Research Paper


1 Department of Biotechnology, Faculty of Advanced Sciences and Technologies, University of Isfahan, Isfahan, IR Iran

2 Department of Biochemistry, School of Pharmacy and Pharmaceutical Sciences, Bioinformatics Research Center, Isfahan University of Medical Sciences, Isfahan, IR Iran

3 Ramin Agricultural and Natural Resources University, Mollasani, Ahwaz, IR Iran



Background: Taxol is one of the most important anti-cancer drugs, which is obtained from yew trees (Taxus sp.). T+he fist step in side chain assembly of taxol is catalyzed by phenylalanine aminomutase, which converts α-phenylalanine to β-phenylalanine. Objectives: In this study, for the fist time, we report on the cloning, preliminary expression and characterization of a full-length gene and cDNA encoding phenylalanine aminomutase from Taxus baccata L. Materials and Methods: Comparison of the full-length gene with other ones identifid from the Taxus species showed high similarity, particularly with Taxus x media. Results: The results showed that the expression level of this gene in Taxus baccata is very low and therefore this enzymatic step could be a rate limiting step in the taxol biosynthesis pathway. Successful amplifiation of the cDNA was only obtained from RNA samples isolated from methyl jasmonate elicited suspension cells of Taxus baccata. The cloned cDNA contained a 2064 bp open reading frame encoding a protein composed of 687 amino acids. Sequence comparison analysis revealed that the gene is very similar (98 - 99 %) with respect to the nucleotide and amino acid sequences in diffrent Taxus species and also share the signature active site motif (175ASG177). Conclusions: The predicted structure of TbPAM was analyzed using bioinformatic tools. The results indicated that the protein has similar overall folding to tyrosine aminomutase.


1. Frense D. Taxanes: perspectives for biotechnological production.
Appl Microbiol Biotechnol. 2007;73(6):1233-40
2. Kai G, Miao Z, Qiu C, Zhang L, Zhao L, Li Z, et al. Molecular cloning and characterization of a taxadienol acetyl transferase cDNA
from Taxus x media. Plant Sci. 2004;167(4):759-64
3. Abbasi Kejani A, Hosseini Tafreshi SA, Khayyam Nekouei SM, Mofi MR. Effient isolation of high quality nucleic acids from different tissues of Taxus baccata L. Mol Biol Rep. 2010;37(2):797-800
4. Walker KD, Klettke K, Akiyama T, Croteau R. Cloning, heterologous expression, and characterization of a phenylalanine
aminomutase involved in Taxol biosynthesis. J Biol Chem.
5. Steele CL, Chen Y, Dougherty BA, Li W, Hofstead S, Lam KS, et al.
Purifiation, cloning, and functional expression of phenylalanine aminomutase: the fist committed step in Taxol side-chain
biosynthesis. Arch Biochem Biophys. 2005;438(1):1-10
6. Ghassempour A, Rezadoost H, Ahmadi M, Aboul-Enein HY.
Seasons Study of Four Important Taxanes and Purifiation of
10-Deacetylbaccatin III from the Needles of Taxus baccata L. by
Two-Dimensional Liquid Chromatography. J Liquid Chromatography Related Tech. 2009;32(10):1434-47
7. Kajani AA, Mofi MR, Abolfazli K, Tafreshi SA. Encapsulated activated charcoal as a potent agent for improving taxane synthesis and recovery from cultures. Biotechnol Appl Biochem.
8. Collins D, Mill RR, Moller M. Species separation of Taxus baccata,
T. canadensis, and T. cuspidata (Taxaceae) and origins of their
reputed hybrids inferred from RAPD and cpDNA data. Am J Bot.2003;90(2):175-82
9. Wang YD, Yuan YJ, Wu JC. Induction studies of methyl jasmonate
and salicylic acid on taxane production in suspension cultures
of Taxus chinensis var. mairei. Biochem Eng J. 2004;19(3):259-65
10. Louie GV, Bowman ME, Moffi MC, Baiga TJ, Moore BS, Noel JP.
Structural determinants and modulation of substrate specifiity in phenylalanine-tyrosine ammonia-lyases. Chem Biol.
11. Montavon TJ, Christianson CV, Festin GM, Shen B, Bruner SD.
Design and characterization of mechanism-based inhibitors
for the tyrosine aminomutase SgTAM. Bioorg Med Chem Lett.
12. Wu B, Szymanski W, Wietzes P, de Wildeman S, Poelarends GJ,
Feringa BL, et al. Enzymatic synthesis of enantiopure alphaand beta-amino acids by phenylalanine aminomutase-catalysed amination of cinnamic acid derivatives. Chembiochem.
13. Christianson CV, Montavon TJ, Festin GM, Cooke HA, Shen B,
Bruner SD. The mechanism of MIO-based aminomutases in betaamino acid biosynthesis. J Am Chem Soc. 2007;129(51):15744-5
14. Dixon RA, Dey PM, Lamb CJ. Phytoalexins: enzymology and molecular biology. Adv Enzymol Relat Areas Mol Biol. 1983;55:1-136