Effect of genetically modified Lactococcus lactis cell-envelope proteinases with altered specificity on the course of casein degradation under cheese conditions

Abstract

Abstract The specificity of the cell-bound, wild-type proteinase (PrtP) of Lactococcus lactis subsp. cremoris SK11 towards the αs1-casein(1–23) fragment under simulated cheese conditions was compared with that of several mutants of PrtP. A unique specificity resulted from substitution of the substrate-binding site residues AKT 137–139 in the wild-type with GLA in the mutant PrtP. This different specificity clearly modifies the breakdown of chymosin-generated primary cheese peptides under cheese conditions. The consequences for amino acid production were investigated with cells which were optimally permeabilized with respect to accessibility and activity of the intracellular proteolytic system. Unlike untreated cells, these cells showed efficient peptide uptake and intracellular conversion (involving peptides of up to 15 residues), similar to that occurring in a normal Gouda cheese. No significant differences were found in the composition of the amino acid pools generated by permeabilized cells containing either the wild-type or the mutant PrtP.