Study on Milk-Clotting Mechanism of Rennet-Like Enzyme from Glutinous Rice Wine: Proteolytic Property and the Cleavage Site on κ-Casein

The SDS-PAGE (15%) profile of purified enzyme from glutinous rice wine. Lane 1 = protein molecular weight markers; line 2 = crude enzyme; lane 3 = purified enzyme.

The SDS-PAGE electrophoregrams of bovine milk proteins hydrolyzed by protease of glutinous rice wine. Lanes 1, 3, 5, 7, 9, and 11 show the intact κ-, β-, and α-CN; β-LG; whole CN; and albumin (10 mg/mL), respectively, and the corresponding digested is shown in lanes 2, 4, 6, 8, 10, and 12. The direction of migration is shown with an arrow.

Peptide PAGE electrophoregrams of CN compositions by rennet-like protease from glutinous rice wine at reaction times 0, 5, 15, 30, 60, 120, 180, and 240min and 12h (left to right). The main peptide is indicated with an arrow; a = α-CN; b = β-CN; c = κ-CN.

Mass spectrum of the rennet digestion of bovine κ-CN. a = electrospray tandem mass spectrometry, Q-TOF2; b and c = matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (Reflex, Bruker, Billerica, MA). m/z = mass:charge ratio; AI = abundance of ion.

Peptide mass fingerprint of κ-CN fragment from in-gel trypsin digestion. m/z = mass:charge ratio; AI = abundance of ion.

Amino acid sequence of κ-CN and peptide fragments detected by electrospray tandem mass spectrometry, Q-TOF2 (shaded and boxed), and matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (underlined; Autoflex, Bruker, Billerica, MA).