摘要
口虾蛄死后,肌肉蛋白降解速度快,对品质影响大。以特异的丝氨酸蛋白酶荧光底物为筛选手段,通过硫酸铵盐析、DEAE-Sepharose离子交换柱层析及Sephacryl S-200凝胶过滤柱层析,从口虾蛄肌肉中分离得到一种丝氨酸蛋白酶。SDS-PAGE电泳结果表明,该酶得到有效纯化,分子质量约为27 ku。底物特异性分析表明,纯化的蛋白酶可能为胰蛋白酶类的丝氨酸蛋白酶,其最适温度37℃,最适pH 8.5。该酶在温度低于40℃、pH5.0~9.5的条件下具有较好的稳定性,在温度低于20℃时仍有较高的活性。研究证实,纯化的丝氨酸蛋白酶在体外对口虾蛄的肌原纤维蛋白具有明显的分解作用。
This study is aimed to investigate the proteases which lead to the softening of mantis shrimp （Squilla oratoria） muscle. By incubating the muscle crude extracts with the proteinaceous inhibitors, the serine proteinase was indi- cated to play a major role in degradation of mantis shrimp muscle proteins. Consequently, a serine proteinase from the muscle of mantis shrimp was purified to homogeneity by ammonium sulfate precipitation and column chromatographies of DEAE-Sepharose and Sephacryl S-200. SDS-PAGE result showed that the molecular mass of purified protein was about 27 ku. The purified enzyme was indicated to be a trypsin-like serine proteinase as it could cleave fluorogenic substrates having Arg or Lys at P1 position, but not chymotrypsin substrate（Suc-Leu-Leu-Val-Tyr-MCA）. The optimum temperature and pH of the purified enzyme were 37 ℃ and 8.5, respectively. This enzyme was stable below 40 ℃ and in the pH range of 5.0-9.5, and showed high enzyme activity as the temperature below 20 ℃, suggesting its potential applications. The in vitro degradation effect of purified serine proteinase on myofibrillar proteins verified its important role during the softening of mantis shrimp muscle.
出处
《中国食品学报》
EI
CAS
CSCD
北大核心
2013年第11期185-190,共6页
Journal of Chinese Institute Of Food Science and Technology
基金
国家自然科学基金(31271838
31301440)
海洋公益性行业科研专项经费项目(201305015-3)
福建省教育厅科技项目(JA11159)
厦门市科技计划项目(3502Z20133020)
关键词
口虾蛄
丝氨酸蛋白酶
纯化
降解
Mantis shrimp
serine proteinase
purification
degradation
