Cloning, nucleotide sequence and expression in Streptomyces lividans and Escherichia coli of pabB from Lactococcus lactis subsp. lactis NCDO 496.
A gene (pabB) encoding the aminase activity of p-aminobenzoate (PABA) synthase in Lactococcus lactis subsp. lactis was cloned in pIJ41 and expressed in Streptomyces lividans strains defective in PABA biosynthesis. Expression of the gene was associated with a 1.2 kb deletion between the aph promoter and the cloning site in pIJ41. Subcloning in pBR322 and expression in Escherichia coli AB3295 of the cloned L. lactis DNA fragment localized the pabB-complementing gene in a 1.9 kb segment. The nucleotide sequence of this segment contained a 1410 bp open reading frame encoding a 470-amino-acid polypeptide of 50937 Da. The deduced amino acid sequence showed substantial similarity to those reported for PabB and TrpE from several organisms. Synonymous codon usage reflected the low G + C content in the genomic DNA of L. lactis subsp. lactis, and therefore differed markedly from the preferred usage in the S. lividans host. The cloned heterologous pabB DNA was expressed in amounts that allowed accumulation of excreted PABA in cultures of S. lividans transformants.
Arhin
F F
FF
Department of Biology, Dalhousie University, Halifax, Nova Scotia, Canada.
Vining
L C
LC
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D003062:Codon; D004269:DNA, Bacterial; D004926:Escherichia coli; D013294:Lactococcus lactis / Q000201:enzymology / Q000235:genetics*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D017386:Sequence Homology, Amino Acid; D013302:Streptomyces; D000637:Transaminases / Q000235:genetics* / Q000378:metabolism
D003062:Codon; D004269:DNA, Bacterial; C022309:PABA synthase; D000637:Transaminases