Proteomic Comparison of Equine and Bovine Milks on Renneting
Résumé
Rennet-induced coagulation of bovine milk is a complex mechanism in which chymosin specifically hydrolyzes kappa-casein, the protein responsible for the stability of the casein micelle. In equine milk, this mechanism is still unclear, and the protein targets of chymosin are unknown. To reveal the proteins involved, the rennetability of equine milk by calf chymosin was examined using gel free and gel based proteomic analysis and compared to bovine milk RP-HPLC analysis of bovine and equine milks showed the release of several peptides following chymosin incubation. The hydrolyses of equine and bovine casein by chymosin were different, and the major peptides produced from equine milk were identified by mass spectrometry as fragments of beta-casein. Using two-dimensional electrophoresis, equine beta-casein was confirmed as the main target of calf chymosin over 24 h at 30 degrees C and pH 6.5. The gel based analysis of equine milk discriminated between the different individual proteins and provided information on the range of isoforms of each protein as a result of post translational modifications, as well as positively identified for the first time several isoforms of kappa-casein. In comparison to bovine milk, kappa-casein isoforms in equine milk were not involved in chymosin-induced coagulation. The intensity of equine beta-casein spots decreased following chymosin addition, but at a slower rate than bovine kappa-casein.