Highly efficient production of the staphylococcal nuclease reporter in Lactobacillus bulgaricus governed by the promoter of the hlbA gene.
Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) genome sequence analysis revealed the presence of two genes that encode histone-like HU proteins (hlbA and hlbB) showing extensive similarity to other bacterial homologues. These genes were found to be extremely conserved among several L. bulgaricus strains. The hlbA gene was shown to be constitutively transcribed from a unique promoter (phlbA) during normal growth, whereas hlbB did not seem to be expressed under usual laboratory conditions. Using a reporter cassette in which the staphylococcal nuclease was fused at its N-terminus to the lactococcal signal peptide Usp45 (SP Usp45), we have demonstrated that phlbA promotes high expression of the reporter in L. bulgaricus, which correlated with an abundant secretion of the mature nuclease in the supernatant fraction. Quantification of the exported enzyme reveals a secretion level approximately threefold higher when the expression of the reporter was under the control of phlbA compared with the lactococcal usp45 promoter. Together, these results indicate that phlbA is suitable for gene expression in L. bulgaricus, that SP Usp45 is functionally recognized and processed by the L. bulgaricus secretion machinery and that the nuclease reporter gene can be used for the identification of exported products in this bacterium.
Chouayekh
Hichem
H
Laboratoire d'Enzymes et de Métabolites des Procaryotes, Centre de Biotechnologie de Sfax, Sfax, Tunisie. hichem.chouayekh@cbs.rnrt.tn
Serror
Pascale
P
Boudebbouze
Samira
S
Maguin
Emmanuelle
E
D001483:Base Sequence; D017930:Genes, Reporter; D048668:Lactobacillus delbrueckii / Q000235:genetics*; D008836:Micrococcal Nuclease / Q000096:biosynthesis* / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics
D011993:Recombinant Fusion Proteins; D008836:Micrococcal Nuclease