Proteolysis of the zona pellucida of mouse ova.

branes, was measured by the method of Abrams [S]. Endopeptidase activity was measured by incubating 50 pl of each fraction with 450 pI of peptide-coumarin derivative, each at 0.1 11 mM in 50 mM-Tris-HCI, pH 7.5. The release of free 7-amino-4-methylcoumarin was quantified fluorimetrically using excitation and emission wavelengths of 370 nm and 440 nm, respectively. The results obtained for S. cremoris HP are presented in Table 1, and similar results were obtained with S. cremoris AM2. Glucosamine was highest in cell walls of the cellular fractions assayed. The highest levels of magnesium ATPase were found in the cell membrane fraction, whereas the highest levels of lactate dehydrogenase were found in cytoplasm. The majority of the endopeptidase activity as measured by each of the peptide-coumarin derivatives employed was observed in the cytoplasmic fractions with significant levels of activity in the cell membrane fractions. Little or no activity against the peptide-coumarin derivatives could be found in the extracellular fluid, wash or cell wall fraction. It is of note that the previously mentioned proteinase activity of cell walls could not be detected by these substrates. Our results indicate that the endopeptidase activities observed using peptide-coumarin derivatives as substrates, being predominantly located in the cytoplasm, are likely to be significant in cheese ripening only on the lysis of the cell.