Overexpression of a designed 2.2 kb gene of eukaryotic phenylalanine ammonia-lyase in Escherichia coli.
Phenylalanine ammonia-lyase (EC 4.3.1.5) is a key enzyme in the secondary metabolism of higher plants catalyzing the non-oxidative conversion of L-phenylalanine into transcinnamate. The nucleotide sequence of its 2.2 kb gene was designed for expression in Escherichia coli and synthesized in a single reaction from 108 oligonucleotides using assembly PCR. After amplification, the gene was cloned into the expression vector pT7-7 and coexpressed with the chaperone HSP-60 system. The expression system yielded 70 mg of fully active enzyme per liter culture.
Baedeker
M
M
Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität, Freiburg im Breisgau, Germany.
Schulz
G E
GE
D019661:Apiaceae / Q000201:enzymology / Q000235:genetics; D001483:Base Sequence; D003062:Codon; D003461:Crystallography; D004926:Escherichia coli / Q000235:genetics; D002479:Inclusion Bodies / Q000235:genetics; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000235:genetics; D010650:Phenylalanine Ammonia-Lyase / Q000235:genetics* / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods
D003062:Codon; D018832:Molecular Chaperones; D009841:Oligonucleotides; D010650:Phenylalanine Ammonia-Lyase