Structural changes and interactions involved in the Ca(2+)-triggered stabilization of the cell-bound cell envelope proteinase in Lactococcus lactis subsp. cremoris SK11.

The cell-bound cell envelope proteinase (CEP) of the mesophilic cheese-starter organism Lactococcus lactis subsp. cremoris SK11 is protected from rapid thermal inactivation at 25 degrees C by calcium bound to weak binding sites. The interactions with calcium are believed to trigger reversible structural rearrangements which are coupled with changes in specific activity (F.

Role of calcium in activity and stability of the Lactococcus lactis cell envelope proteinase.

The mature lactococcal cell envelope proteinase (CEP) consists of an N-terminal subtilisin-like proteinase domain and a large C-terminal extension of unknown function whose far end anchors the molecule in the cell envelope. Different types of CEP can be distinguished on the basis of specificity and amino acid sequence. Removal of weakly bound Ca2+ from the native cell-bound CEP of Lactococcus lactis SK11 (type III specificity) is coupled with a significant reversible decrease in specific activity and a dramatic reversible reduction in thermal stability, as a result of which no activity at 25 degrees C (pH 6.

The occurrence of two intracellular oligoendopeptidases in Lactococcus lactis and their significance for peptide conversion in cheese.

Two intracellular oligopeptide-preferring endopeptidases have been detected in Lactococcus lactis. A neutral thermolysin-like oligoendopeptidase (NOP) has been purified to homogeneity and an alkaline oligoendopeptidase has been partially purified. The specificity of the oligoendopeptidases towards important intermediary cheese peptides, produced by chymosin action on the caseins, clearly differs from that of the cell-envelope proteinase (CEP).

Purification and Characterization of Cystathionine (beta)-Lyase from Lactococcus lactis subsp. cremoris B78 and Its Possible Role in Flavor Development in Cheese.

An enzyme that degrades sulfur-containing amino acids was purified from Lactococcus lactis subsp. cremoris B78; this strain was isolated from a mixed-strain, mesophilic starter culture used for the production of Gouda cheese. The enzyme has features of a cystathionine (beta)-lyase (EC 4.

Evidence for a large dispensable segment in the subtilisin-like catalytic domain of the Lactococcus lactis cell-envelope proteinase.

The Lactococcus lactis SK11 cell-envelope proteinase contains various inserts, located in external loops of the catalytic domain compared with related subtilisins. In this study, protein engineering was employed to determine the function of the largest loop insertion (residues 238-388) relative to the subtilisin structure. By site-directed mutagenesis we have deleted the fragment of the proteinase gene encoding these 151 residues and analyzed the mutant delta 238-388 proteinase for activity, (auto)processing and cleavage specificity.