Cloning and expression of a phenylalanine ammonia-lyase gene (BoPAL2) from Bambusa oldhamii in Escherichia coli and Pichia pastoris

Abstract

Phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) is the first committed enzyme of phenylpropanoid pathway. A PAL gene, designated as BoPAL2, was cloned from a Bambusa oldhamii cDNA library. The open reading frame of BoPAL2 was 2142bp in size encoding a 713-amino acid polypeptide. BoPAL2 was heterologous expressed in Escherichia coli and Pichia pastoris. The recombinant proteins were exhibited PAL and tyrosine ammonia-lyase activities. The recombinant BoPAL2 had a subunit mass of 80kDa and existed as a homotetramer. The optimum temperature and pH of BoPAL2 were 50-60 degrees C and 8.5-9.0, respectively. The K(m) and k(cat) values of BoPAL2 expressed in E. coli were 250microM and 10.12s(-1). The K(m) and k(cat) values of BoPAL2 expressed in P. pastoris were 331microM and 16.04s(-1). The recombinant proteins had similar biochemical properties and kinetic parameters with PALs reported in other plants.