Expression and Activity Assay of Recombinant Phenylalanine Ammonia Lyase of Glycine max L. in E.coli

Abstract

Abstract: Phenylalanine ammonia-lyase(PAL) gene of Glycine max L (GM) was cloned by SOE-PCR method from the genomic DNA. The full length exon of PAL expression vector pET-GMPAL was constructed and expressed in E.coli. Catalytic activity of conversion L-phenylalanine into cinnamic acid by the recombination GMPAL is achieved at high specific activity, up to 211.7μmol/g pro·min (3529μkat/kg), higher than the parsley recombinant PAL, and Rhotodotorula glutins PAL. Molecular weight of the expressed PAL protein was estimated to be 77.9kD. The amounts of PAL protein expression level was 8% of total E.coli soluble proteins by SDS-PAGE. Results indicated that soybean recombinant PAL is efficiently expressed in E.coli as fully activated functional enzyme.

Key words: , phenylalanine ammonia lyase (PAL)； pET-31b(+)； gene cloning； recombinant expression； cinnamic acid； splicing by overlap extention(SOE)； polymerase chain reaction(PCR)；

SONG Jian, MA Hui-Qin, HAO Jia, Ren-Fa-Zheng, Chen-Shang-Wu. Expression and Activity Assay of Recombinant Phenylalanine Ammonia Lyase of Glycine max L. in E.coli[J]. FOOD SCIENCE, 2006, 27(7): 29-35.

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Expression and Activity Assay of Recombinant Phenylalanine Ammonia Lyase of Glycine max L. in E.coli

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