Physico-chemical characterization of a milk-clotting fraction extracted from turkey (Meleagris gallopavo) proventriculus

Abstract Valorization of avian by-products such as turkey ( Meleagris gallopavo ) proventriculus in the preparation of milk clotting enzymes constitutes an alternative to commercial rennet. The clotting enzyme of Meleagris gallopavo has been prepared by ammonium sulfate at 50% saturation followed by purification by cation-exchange fast protein liquid chromatography (FPLC), and determination of the clotting protease activity performed on milk incubated at 35 °C. The clotting fractions are concentrated and dialyzed against Tris/HCl pH 7.0 through a membrane with cutoff 8000 Da, and separated by anion-exchange FPLC. The coagulating fractions obtained by cation-exchange chromatography are also separated by reversed-phase high-performance liquid chromatography, which revealed only one fraction able to coagulate milk. Electrophoresis analysis revealed a band with apparent molecular mass of 36.5 kDa and seemed to correspond to pepsin-like enzyme according to primary sequence alignment analyses. Maximal clotting activity was obtained for optimal conditions of temperature of 55 °C and pH 5.4.