Expression of bacillar glutamyl endopeptidase genes in Bacillus subtilis by a new mobilizable single-replicon vector pLF.

The pLF1311 natural plasmid from Lactobacillus fermentum 1311 was used to construct a single-replicon vector suitable for rapid cloning in a wide range of gram-positive hosts and Escherichia coli. The new vector is capable of conjugative mobilization from E. coli to various hosts by conjugal transfer. The final vector (3.4 kb) showed a high segregational and structural stability and a high copy number. Glutamyl endopeptidase genes from Bacillus licheniformis (gseBL) and B. intermedius (gseBI) were cloned in both pLF9 and pLF14 vectors and introduced to B. subtilis. The yield of enzymes in the pLF-derived producers was 6- to 30-fold more than in the natural producers and reached 100-150 mg/L of mature protease.