Milk-clotting activity of enzyme extracts from sunflower and albizia seeds and specific hydrolysis of bovine κ-casein

Milk-clotting activity found in ammonium sulfate-precipitated protein extracts from Albizia lebbeck and Helianthus annuus seeds was studied. Specific clotting activity of albizia seed extract was 15 times higher than that of sunflower seed extract. Zymogram analysis revealed several proteolytic bands in albizia seed extract and one diffuse proteolytic band for sunflower seed extract. Whole bovine casein was incubated with the plant seed extracts or chymosin and some breakdown products were characterized by reversed-phase high-performance liquid chromatography and electrophoresis. Similar to chymosin, the two seed extracts exhibited proteolytic activity toward κ-casein, αs-casein and β-casein, with the highest activity observed for the albizia seed extract. Mass spectrometry analysis showed that the sunflower extract hydrolyzed κ-casein at the Phe105–Met106 bond, as does chymosin. The albizia extract also displayed activity on κ-casein, but the Lys116–Thr117 bond was its preferred target.