The Major Lactococcal Cell Wall Autolysin AcmA Does Not Determine the Rate of Autolysis of Lactococcus lactis subsp. cremoris 2250 in Cheddar Cheese

Abstract

Abstract A range in the levels of autolysis of 17 commercial Lactococcus lactis cheese starters was found after growth and incubation in milk for 72 h at 30°C. Autolysis levels in milk usually (but not always) correlated with autolysis in 0.1 m Na phosphate buffer. One strain, Lc. lactis subsp. cremoris strain 2250, was highly autolytic. The major lactococcal autolysin (N-acetyl muramidase; AcmA) was cloned from 2250 and sequenced. Comparison of the predicted amino acid sequence with AcmA from Lc. lactis MG1363 showed the presence of ten mostly conserved substitutions, equivalent to 98% amino acid homology. Three substitutions (all conserved) were in the active site domain, but none of these involve the acidic amino acids expected to be required for muramidase activity. This concurs with earlier results (Riepe et al. (1997) Applied and Environmental Microbiology 63 , 3757–3763) showing that there is no significant difference in enzymatic activities of the two AcmAs. A 2250 acmA deletion mutant (2250Δ acmA ), completely lacking any AcmA activity, was used to assess the contribution of AcmA to the autolysis of 2250 in milk and Cheddar cheese. In milk, AcmA contributed significantly to high autolysis of 2250. Nevertheless, autolysis of 2250Δ acmA in milk was higher than most of 16 other Lc. lactis strains, all of which have AcmA present. By contrast, in cheese, 2250Δ acmA autolysed almost to the same extent as 2250 wild-type. Factors other than AcmA must therefore cause the high levels of autolysis seen in Lc. lactis subsp. cremoris 2250 in cheese. These factors may be important in causing high autolysis of other autolytic Lc. lactis strains.