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Characterization of a Serine Proteinase of Several Enterococcus Durans and Enterococcus Faecium Strains
Abstract
A study of casein hydrolysis by Enterococcus durans 19432 and 11576, and Enterococcus faecium 19434 and 3 5667 was completed. These strains of E. durans and E. faecium are used as starter cultures in some artisanal cheeses. Casein-degradation was demonstrated by changes in patterns seen on SDS-PAGE. Strains of the enterococci studied did not possess a cell envelope proteinase when purified casein preparations were incubated with the supernatant from cells treated in calcium-free phosphate buffer. Intracellular proteinase activity was shown by incubating purified casein preparations with whole cell suspensions. Based on Polymerase Chain Reaction amplification experiments using total cellular DNA preparations and specific primers for the serine proteinase gene (PrtP) from Lactococcus lactis ssp. cremoris SK 11, it was inferred that E. durans and E. faecium strains possess a similar serine proteinase gene. Ability to hydrolyze casein fractions (αs-, β-, and κ-casein) varied considerably. This was attributed to strain and species differences. Intracellular proteinase activity was detected in all four strains. Complete β-casein hydrolysis by all the Enterococcus strains tested was detected within 72 hours. Enterococcus durans 11576 did not hydrolyze αs -casein within 72 hours. Enterococcus durans 19432, E. faecium 19434 and E. faecium 35667 hydrolyzed both αs - and β-casein and would be considered to have PIii type proteinase activity. Enterococcus durans 11576 hydrolyzed β-casein only and would be considered to have PI type proteinase. Enterococcus durans 11576, E. faecium 19434 and 35667 appeared to be plasmid free; this suggested that the proteinase genes in these strains are chromosomally located as compared with plasmid location for the cell envelope proteinase of L. lactis ssp. cremoris SK 11.