title: Vector-mediated chromosomal integration of the glutamate decarboxylase gene in Streptococcus thermophilus.
abstract: The integrative vector, pINTRS, was used to transfer glutamate decarboxylase (GAD) activity to Streptococcus thermophilus ST128 thereby allowing for the production of γ-aminobutyric acid (GABA). In pINTRS, the gene encoding glutamate decarboxylase, gadB, was flanked by DNA fragments homologous to a S. thermophilus pseudogene to allow for integration at a non-essential locus on the chromosome. Screening techniques confirmed the insertion of gadB with either its endogenous promoter or the S. thermophilus P2201 promoter, resulting in the generation of recombinant strains, ST128/gadB or ST128/P2201-gadB. Following the integration event unwanted plasmid DNA, specifically the erythromycin resistance gene, was eliminated from the recombinant strains. Based on the production of GABA, activities of GAD for ST128/gadB and ST128/P2201-gadB were 30.6 ± 6 and 27.9 ± 7.2 μM/mg dry cell wt, respectively.
authors: Eastern Regional Research Center, Agricultural Research Service, United States Department of Agriculture, Wyndmoor, PA 19038, USA. john.renye@ars.usda.gov
authors: John A
authors: JA
authors: Renye
authors: George A
authors: GA
authors: Somkuti
mesh_terms: D005822:Genetic Vectors; D005968:Glutamate Decarboxylase / Q000235:genetics* / Q000378:metabolism*; D060847:Metabolic Engineering; D016254:Mutagenesis, Insertional; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D011995:Recombination, Genetic; D048249:Streptococcus thermophilus / Q000235:genetics* / Q000378:metabolism*; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D005680:gamma-Aminobutyric Acid; D005968:Glutamate Decarboxylase