Characterization of transcription initiation and termination signals of the proteinase genes of Lactococcus lactis Wg2 and enhancement of proteolysis in L. lactis

The transcription initiation signals of the prtP and prtM genes specifying the proteolytic activity of Lactococcus lactis subsp. cremoris Wg2 were mapped by primer extension. The strength of these promoters was analyzed with promoter-screening vector pGKV410, and they appeared to be weaker than previously isolated promoters of strain Wg2. In addition, a putative transcription terminator downstream of the prtP gene was characterized by using the terminator-screening vector pGKV259. The putative terminator decreased the transcription activity of lactococcal promoter P59 by approximately 70% in both Bacillus subtilis and L. lactis. Deletion of a part of the stem-loop structure of the terminator decreased the negative effect on transcription, indicating that the structure could indeed function as a terminator of transcription. The proteolytic activity of the lactococcal host was enhanced by placing the originally oppositely oriented prt genes in tandem and replacing the relatively weak promoters upstream of the prt genes with the stronger promoter, P32, from the chromosome of L. lactis Wg2.