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Appl. Environ. Microbiol., Oct 1997, 3757-3763, Vol 63, No. 10
HR Riepe, CJ Pillidge, PK Gopal and LL Mckay
Two highly autolytic Lactococcus lactis subsp. cremoris strains (CO and
2250) were selected and analyzed for their autolytic properties. Both
strains showed maximum lysis when grown in M17 broth containing a limiting
concentration of glucose (0.4 to 0.5%) as the carbohydrate source. Lysis
did not vary greatly with pH or temperature but was reduced when strains
were grown on lactose or galactose. Growth in M17 containing excess glucose
(1%) prevented autolysis, although rapid lysis of L. lactis subsp. cremoris
CO did occur in the presence of 1% glucose if sodium fluoride (an inhibitor
of glycolysis) was added to the medium. Maximum cell lysis in a buffer
system was observed early in the stationary phase, and for CO, two pH
optima were observed for log-phase and stationary-phase cells (6.5 and 8.5,
respectively). Autolysins were extracted from the cell wall fraction of
each strain by using either 4% sodium dodecyl sulfate (SDS), 6 M guanidine
hydrochloride, or 4 M lithium chloride, and their activities were analyzed
by renaturing SDS-polyacrylamide gel electrophoresis on gels containing
Micrococcus luteus or L. lactis subsp. cremoris CO cells as the substrate.
More than one lytic band was observed on each substrate, with the major
band having an apparent molecular mass of 48 kDa for CO. Each lytic band
was present throughout growth and lysis. These results suggest that at
least two different autolytic enzymes are present in the autolytic L.
lactis subsp. cremoris strains. The presence of the lactococcal cell wall
hydrolase gene, acmA (G. Buist, J. Kok, K. J. Leenhouts, M. Dabrowska, G.
Venema, and A. J. Haandrikman, J. Bacteriol. 177:1554-1563, 1995), in
strains 2250 and CO was confirmed by Southern hybridization. Analysis of an
acmA deletion mutant of 2250 confirmed that the gene was involved in cell
separation and had a role in cell lysis.
Copyright © 1997, American Society for Microbiology
Characterization of the Highly Autolytic Lactococcus lactis subsp. cremoris Strains CO and 2250
Department of Food Science and Nutrition, University of Minnesota, St. Paul, Minnesota 55108, and New Zealand Dairy Research Institute, Palmerston North, New Zealand
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