The major lactococcal cell wall autolysin AcmA does not determine the rateof autolysis of Lactococcus lactis subsp. cremoris 2250 in cheddar cheese

A range in the levels of autolysis of 17 commercial Lactococcus lactis chee se starters was found after growth and incubation in milk for 72 h at 30 de grees C. Autolysis levels in milk usually (but not always) correlated with autolysis in 0.1 M Na phosphate buffer. One strain, Lc. lactis subsp. cremo ris strain 2250, was highly autolytic. The major lactococcal autolysin (N-a cetyl muramidase; AcmA) was cloned from 2250 and sequenced. Comparison of t he predicted amino acid sequence with AcmA from Lc. lactis MG1363 showed th e presence of ten mostly conserved substitutions, equivalent to 98% amino a cid homology. Three substitutions (all conserved) were in the active site d omain, but none of these involve the acidic amino acids expected to be requ ired for muramidase activity. This concurs with earlier results (Riepe et a l. (1997) Applied and Environmental Microbiology 63, 3757-3763) showing tha t there is no significant difference in enzymatic activities of the two Acm As. A 2250 acmA deletion mutant (2250 Delta acmA), completely lacking any A cmA 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 2250 Delta acmA in m ilk was higher than most of 16 other Lc. lactis strains, all of which have AcmA present. By contrast, in cheese, 2250 Delta acmA autolysed almost to t he same extent as 2250 wild-type. Factors other than AcmA must therefore ca use 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. (C) 1999 Elsevier Science Ltd. All rights res erved.