CLONING OF PROMOTER-LIKE SEQUENCES FROM LACTOBACILLUS-PARACASEI SUBSPPARACASEI CG11 AND THEIR EXPRESSION IN ESCHERICHIA-COLI, LACTOCOCCUS-LACTIS, AND LACTOBACILLUS-REUTERI

Fragments of chromosomal DNA from Lactobacillus paracasei subsp. parac asei CG11 (formerly Lactobacillus casei CG11) capable of functioning a s promoters were isolated using the broad host range, promoter-probe v ector pGKV210. Five such fragments designated P61, P79, P80, P116, and P144 were completely sequenced and analyzed. Fragment P61 had the hig hest transcriptional efficiency in Escherichia coil and Lactobacillus reuteri whereas P80 was the most active in Lactococcus lactis. In gene ral, the orders of the transcriptional strengths were almost identical in E. coil and Lactobacillus reuteri but different from that in Lacto coccus lactis. Mapping of the 5' end of cat mRNA showed that different regions of fragments P79 and P144 were used as promoters in Lactococc us lactis than in E. coil and Lactobacillus reuteri. Analysis of these DNA sequences revealed that the putative -35 and -10 hexanucleotides resembled those of E. coli, Bacillus subtilis, and lactococci. The spa cing between these two hexanucleotides and between the putative -10 he xanucleotide and the transcriptional start point (A residues predomina ted) ranged from 17 to 18 base pairs and from 5 to 7 base pairs, respe ctively. Each of the cloned Lactobacillus paracasei CG11 promoter-like fragments contained an AT-rich sequence upstream of the putative -35 region (from 60 to 73%).