title: Proteinase overproduction in Lactococcus lactis strains: regulation and effect on growth and acidification in milk.
abstract: Multicopy plasmids that contained the complete of 3'-deleted forms of the proteinase (prtP) gene of Lactococcus lactis subsp. cremoris SK11 under the control of different promoters were constructed and introduced into Prt- lactococcal strains. The production and location of the SK11 proteinase was determined in different hosts grown in industrial and laboratory media. In spite of the 10-fold-higher copy number of the prt genes, no overproduction of proteinase was observed in strain SK1128, a Prt- derivative of L. lactis subsp. cremoris SK112. In contrast, an approximately threefold overproduction of the cell envelope-located or fully secreted proteinase was found in strain MG1820 compared with that of its parental strain L. lactis subsp. lactis SH4109. In all strains proteinase production appeared to be regulated by the medium composition. Highest proteinase production of the SK11 derivatives was found in milk, in contrast to derivatives of SH4109 that produced most proteinase in whey permeate medium. Analysis of single strains with different levels of proteinase production or mixed cultures containing various ratios of Prt+ and Prt- cells indicated that the amount of proteinase produced per cell or culture determines the specific growth rate in milk. Overproduction of cell envelope-located or secreted proteinase in strain MG1820 resulted in a 20%-higher specific growth and acidification rate in milk compared with that in the wild-type strain SH4109. These results indicate that the growth of lactococci in milk is limited by the caseinolytic activity of the proteinase.
authors.affiliation: Molecular Genetics Group, Department of Biophysical Chemistry, Netherlands Institute for Dairy Research (NIZO).
authors.forename: P G
authors.initials: PG
authors.lastname: Bruinenberg
authors.forename: P
authors.initials: P
authors.lastname: Vos
authors.forename: W M
authors.initials: WM
authors.lastname: De Vos
mesh_terms: D000818:Animals; D001483:Base Sequence; D004269:DNA, Bacterial; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000096:biosynthesis* / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D006863:Hydrogen-Ion Concentration; D013294:Lactococcus lactis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D008892:Milk / Q000382:microbiology*; D008969:Molecular Sequence Data; D010957:Plasmids; D015183:Restriction Mapping
chemical_list: D004269:DNA, Bacterial; D010450:Endopeptidases