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The pLF1311 natural plasmid from Lactobacillus fermentum1311 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. colito 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.

A 15‐bp mini‐gene was introduced into Bacillus subtilisand into stable protoplast‐like L‐forms of Proteus mirabilis. This mini‐gene encoded the peptide MVLFV and modeled a fragment of Escherichia coli23S rRNA responsible for E. colierythromycin (Ery) resistance. Expression of the introduced mini‐gene conferred permanent Ery resistance on B. subtilis. In L‐forms of P. mirabilis, the Ery‐protective effect was maintained in the course of several generations. Herewith, the mechanism of Ery resistance mediated by expression of specific short peptides was shown to exist in evolutionary distant bacteria. Three new plasmids were constructed containing the gene under study transcriptionally fused with the genes encoding glutamylendopeptidase of Bacillus licheniformisor δ‐endotoxin of Bacillus thuringiensis. The Ery resistance pentapeptide (E‐peptide) mini‐gene served as an efficient direct transcriptional reporter and allowed to select bacillar glutamylendopeptidase with improved productivity. The mini‐genes encoding E‐peptides may be applied as selective markers to transform both Gram‐positive and Gram‐negative bacteria. The small size of the E‐peptide mini‐genes makes them attractive selective markers for vector construction.

A metalloprotease gene of Brevibacillus brevis(npr) was expressed in Escherichia coliin a soluble form as native Npr precursor. A significant fraction of the precursor was spontaneously processed, producing the N‐terminal propeptide and the mature enzyme. A strong inhibition of the mature Npr by its own propeptide in the crude lysate was observed even in the absence of the covalent linkage between them. Pure precursor, propeptide and the mature Npr were isolated and kinetic parameters of the mature enzyme inhibition by the propeptide were determined. The inhibition is of the tight‐binding competitive type with Ki0.17 nM. Inhibition of metalloproteases from Brevibacillus megateriumand thermolysine by the heterologous propeptide of the Npr from B. breviswas much weaker or none.

A 15-bp mini-gene was introduced into Bacillus subtilis and into stable protoplast-like L-forms of Proteus mirabilis. This mini-gene encoded the peptide MVLFV and modeled a fragment of Escherichia coli 23S rRNA responsible for E. coli erythromycin (Ery) resistance. Expression of the introduced mini-gene conferred permanent Ery resistance on B. subtilis. In L-forms of P. mirabilis, the Ery-protective effect was maintained in the course of several generations. Herewith, the mechanism of Ery resistance mediated by expression of specific short peptides was shown to exist in evolutionary distant bacteria. Three new plasmids were constructed containing the gene under study transcriptionally fused with the genes encoding glutamylendopeptidase of Bacillus licheniformis or δ-endotoxin of Bacillus thuringiensis. The Ery resistance pentapeptide (E-peptide) mini-gene served as an efficient direct transcriptional reporter and allowed to select bacillar glutamylendopeptidase with improved productivity. The mini-genes encoding E-peptides may be applied as selective markers to transform both Gram-positive and Gram-negative bacteria. The small size of the E-peptide mini-genes makes them attractive selective markers for vector construction.