Milk-clotting mechanism of Dregea sinensis Hemsl. protease.
Milk-clotting mechanism of Dregea sinensis Hemsl. protease.
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DOI：
10.3168/jds.2015-9851
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发表时间：
2015-12
影响因子：
3.5
通讯作者：
Ya-li Zhang;Hongyan Wang;Liang Tao;Aixiang Huang
中科院分区：
文献类型：
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作者：
Ya-li Zhang;Hongyan Wang;Liang Tao;Aixiang Huang
山黄麻在中国云南，被用作牛奶凝固剂来生产羊奶蛋糕。然而，凝乳化合物的组成和相关的机制还没有报道。从茎中提取粗蛋白酶，纯化，然后用Millipore超滤离心管分离。通过电喷雾质谱鉴定，半胱氨酸蛋白酶（procerain B）为主要的凝乳蛋白，其分子量为23.8 kDa。该蛋白酶可部分降解α-酪蛋白（CN），完全降解β-和κ-CN，κ-CN降解导致凝乳。通过基质辅助激光解吸/电离飞行时间质谱法和肽测序仪，分别确定的肽馏分的分子量和AA序列。该酶在Ala 90-Gln 91处切割κ-CN，并产生分子量分别为12和6.9 kDa的副κ-CN和酪蛋白巨肽。该切割位点不同于在Phe 105-Met 106处切割的大多数凝乳酶。
Dregea sinensis Hemsl. is used as a milk coagulant to produce goat milk cakes in Yunnan, China. However, the composition of milk-clotting compounds and the related mechanism have not been reported. Crude protease was extracted from the stem, purified, and then separated with a Millipore ultrafiltration centrifuge tube. Cysteine protease (procerain B) was identified as the main milk-clotting protein through electrospray ionization mass spectrometry, and its molecular weight was 23.8 kDa. The protease can partially degrade α-casein (CN) and completely degrade β- and κ-CN, and κ-CN degradation resulted in milk clotting. The molecular weight and AA sequence of the peptide fractions were determined through matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and a peptide sequencer, respectively. The enzyme cleaved κ-CN at Ala90-Gln91 and produced deputy κ-CN and caseinomacropeptide with molecular weights of 12 and 6.9 kDa, respectively. This cleavage site differed from the majority of chymosins cleaved at Phe105-Met106.