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 ␤-lyase (EC 4.4.1.8), a pyridoxal-5-phosphate-dependent enzyme involved in the biosynthesis of methionine and catalyzing an ␣,␤-elimination reaction. It is able to catalyze an ␣,␥-elimination reaction as well, which in the case of methionine, results in the production of methanethiol, a putative precursor of important flavor compounds in cheese. The native enzyme has a molecular mass of approximately 130 to 165 kDa and consists of four identical subunits of 35 to 40 kDa. The enzyme is relatively thermostable and has a pH optimum for activity around 8.0; it is still active under cheese-ripening conditions, viz., pH 5.2 to 5.4 and 4% (wt/vol) NaCl. A possible essential role of the enzyme in flavor development in cheese is suggested.

Partially purified cell wall proteinases of eight strains of Streptococcus cremoris were compared in their action on bovine asl-, 13-, and K-casein, as visualized by starch gel electrophoresis, sodium dodecyl sulfatepolyacrylamide gel electrophoresis, and thin-layer chromatography. Characteristic degradation profiles could be distinguished, from which the occurrence of two proteinases, represented by strain HP and strain AM1, was concluded. The action of the HP-type proteinase PI (also detectable in strains Wg2, C13, and TR) was established by electrophoretic methods to be directed preferentially towards 13-casein. The AM,-type proteinase Pm (also detectable in strain SK11) was also able to degrade 1-casein, but at the same time split asl-and K-casein more extensively than did PI. Strain FD27 exhibited mainly PI activity but also detectable Pm degradation characteristics. The cell wall proteinase preparation of strain E8 showed low PI as well as low Pm activity. All proteinase preparations produced from K-casein positively charged degradation products with electrophoretic mobilities similar to those of degradation products released by the action of the milk-clotting enzyme chymosin. The differences between PI and Pm in mode of action, as detected by gel electrophoresis and thin-layer chromatography, were reflected by the courses of the initial degradation of methyl-14C-labeled (-casein and by the effect of asl-plus K-casein on these degradations. The results are discussed in the light of previous comparative studies of cell wall proteinases in strains of S. cremoris and with respect to the growth of this organism in milk.

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The cell-wall-associated proteolytic systems of several Streptococcus cremoris strains were analyzed by crossed immunoelectrophoresis. At least four immunologically different components of the proteolytic system's were found. One of these proteins was produced by all strains tested. The proteolytic activity of this enzyme was demonstrated with a zymogram staining technique which is based on the degradation of Coomassie-brilliantblue-stainable casein. The crossed-immunoelectrophoresis patterns of the proteolytic systems of different S. cremoris strains indicated that each strain produces a characteristic combination of proteins. On the basis of these combinations, the different S. cremoris strains were classified into four groups.