Characterization of the Cell Wall Proteinase P III of Lactococcus lactis subsp. cremoris Strain AM 1, and its Relationship with the Catalytically Different Cell Wall Proteinase P I/P II of Strain HP

Abstract

Summary The cell wall proteinase PIII of Lactococcus lactis subsp. cremoris AM1 has been purified. In its complexity this proteinase (component 0) shows a remarkable similarity to the catalytically different cell wall proteinase PII (component 1) of L. lactis subsp. cremoris HP. Both proteinases appear as a 135 kDa component in sodiumdodecylsulfate-polyacrylamide gels, but could be converted efficiently into mainly a 80 kDa proteinase and, in addition, a 60 kDa component only under specific, for that matter, completely inactivating thermal conditions. If an otherwise very slow conversion took place at relatively low temperatures, the unstable 80 kDa proteinase remained catalytically active. It is thus responsible for its own degradation. Proteolytically active derivatives with a molecular weight between 135 and 80 kDa, always detectable in the purified proteinase fraction, might either be the result of this action or of an unimolecular action of the 135 kDa proteinase. A higher thermostability of the proteinase fraction of strain HP is due to the more stable, non-converting 125 kDa-component 2 (= PI). Differences in binding and catalytic properties between PI(PII) and PIII can be ascribed to differences in the charge characterictics of two genetically related enzymes. The possible nature of the 135 kDa proteinase is discussed and, taking into account all the available biochemical and genetical information, a working hypothesis describing the structural integration of the proteinase within the cell envelope is presented. This model postulates the involvement of a transmembrane integral, anchor protein unit which binds the proteinase by Ca++-dependent interactions.