In situ proteolytic activity of Lactobacillus helveticus and stretchability of Swiss-Type Cheese

In contrast to other lactic acid bacteria, Lactobacillus helveticus can possess two cell-envelope proteinases (CEPs) called PrtH2 and PrtH (Genay et al 2009, Appl. Env. Microbiol.. 75 (10), 3238-3249; Sadat et al 2011a, in press Int. J. Food Microbiol.). These CEPs exhibit different cleavage specificity on pure s1 -casein (Sadat-Mekmene et al., 2011b, Appl. Env. Microbiol. 77(1) 179-186). The aim of this work was to investigate the proteolytic activity of L. helveticus strains in cheese matrix according to the number of their CEPs and the cheese stretchability. Two strains were selected, ITGLH77 and ITGLH1 which possess one CEP, PrtH2, and two CEPs, PrtH and PrtH2, respectively. The proteolysis was monitored during ripening:i) casein hydrolysis by urea-PAGE; ii) peptide pattern of the cheese aqueous extracts first by SDS-PAGE and second by RP-HPLC. Three chromatographic fractions were collected and peptides identified by RP-HPLC coupled on-line with tandem mass spectrometry. In parallel, the dynamic of stretchability of Emmental cheese was measured in order to determine the contribution of CEPs in its generation, since L. helveticus is known to enhance stretching properties in contrast to other lactobacilli, such as L. delbrueckii subsp. lactis . The microstructure of the strands of Emmental cheeses was observed by confocal laser scanning microscopy. The stretchability of Emmental was significantly higher in cheese manufactured with strain ITGLH77. By using Principal Component Analyses, the stretching properties were found to be correlated with the presence of peculiar peptides derived from primary proteolysis, hydrophobic peptides, containing more than 20 amino acid residues and derived from s1 -casein, which were accumulated in cheese manufactured using strain ITGLH77. The presence of these peptides suggest their less hydrolysis by this strain, in agreement with results observed in vitro (Sadat-Mekmene et al., 2011b), namely different cleavage specificity of CEPs PrtH et PrtH2 on pure s1 -casein