Molecular characterization of an inducible p-coumaric acid decarboxylase from Lactobacillus plantarum: gene cloning, transcriptional analysis, overexpression in Escherichia coli, purification, and characterization

6533b825fe1ef96bd1281fa9

RESEARCH PRODUCT

Molecular characterization of an inducible p-coumaric acid decarboxylase from Lactobacillus plantarum: gene cloning, transcriptional analysis, overexpression in Escherichia coli, purification, and characterization

Charles Diviès Lise Barthelmebs Jean-françois Cavin

subject

Transcription Genetic Carboxy-Lyases Molecular Sequence Data macromolecular substances Molecular cloning medicine.disease_cause Polymerase Chain Reaction Applied Microbiology and Biotechnology Open Reading Frames Lactococcus Gene expression Escherichia coli medicine Genomic library Amino Acid Sequence Cloning Molecular Promoter Regions Genetic Escherichia coli Gene Gene Library Recombination Genetic Electronic Data Processing Base Sequence Ecology biology Nucleic acid sequence Chromosome Mapping Nucleic Acid Hybridization hemic and immune systems Gene Expression Regulation Bacterial Blotting Northern biology.organism_classification Molecular biology Recombinant Proteins Blotting Southern Lactobacillus RNA Bacterial Terminator (genetics)Biochemistry Enzyme Induction Electrophoresis Polyacrylamide Gel Lactobacillus plantarum Research Article Food Science Biotechnology

description

By using degenerate primers designed from the first 19 N-terminal amino acids of Lactobacillus plantarum p-coumaric acid decarboxylase (PDC), a 56-bp fragment was amplified from L. plantarum in PCRs and used as a probe for screening an L. plantarum genomic bank. Of the 2,880 clones in the genomic bank, one was isolated by colony hybridization and contained a 519-bp open reading frame (pdc gene) followed by a putative terminator structure. The pdc gene is expressed on a monocistronic transcriptional unit, which is transcribed from promoter sequences homologous to Lactococcus promoter sequences. No mRNA from pdc and no PDC activity were detected in uninduced cell extracts, indicating that the expression is transcriptionally regulated by p-coumaric acid, which corresponds to an activation factor up to 6,000. The pdc gene was overexpressed constitutively in Escherichia coli, and the recombinant enzyme was purified and characterized.

year	journal	country	edition	language
1997-05-01	Applied and Environmental Microbiology

https://doi.org/10.1128/aem.63.5.1939-1944.1997