Proteolytic activity of three variants of fermentation-produced chymosin on sodium caseinate and during Cheddar cheese ripening

Proteolytic activities and specificities of three types of fermentation-produced chymosin were investigated in cheese and in sodium caseinate (NaCN) digests made using these coagulants. The highest level of enzyme required to hydrolyse all intact αS₁-CN in NaCN solution at pH 5.2 over 24 h was for a modified camel chymosin (mCC; 13.34 IMCU mL⁻¹), followed by camel chymosin (4 IMCU mL⁻¹) and bovine chymosin (0.4 IMCU mL⁻¹). Many peptides were identified using liquid chromatography-mass spectrometry from both Cheddar cheese and NaCN digests produced using each chymosin. Besides previously reported casein-derived peptides produced by bovine and camel chymosins and corresponding cleavage sites, several new cleavage sites were identified. The proteolytic specificity of mCC on casein was determined. Most of the peptides observed in Cheddar cheese samples were also identified in NaCN digests. The overall proteolytic activity of mCC was the lowest, which may have implications for ripening and functionality of cheese.