Kammoun Maher, Gargouri Youssef, Bezzine Sofiane
Laboratoire de Biochimie et de Génie Enzymatique des Lipases, ENIS, Sfax, Tunisia.
Pancreas. 2008 Apr;36(3):302-8. doi: 10.1097/MPA.0b013e31814b9751.
The aim of the present study was to purify and characterize classic pancreatic lipase from the reptile turtle (TuPL).
The lipase was purified from the fresh pancreas extract followed by diethylamino ethyl-cellulose chromatography, Sephacryl S-200 gel filtration, and a Mono-Q Sepharose chromatography.
Turtle pancreatic lipase is a serine enzyme and it contains only 1 free cysteine. Its activity is maximum at pH 8.2 and 37 degrees C. A specific activity of 10.000 U/mg and 5.000 U/mg were measured titrimetrically on tributyrin and olive oil emulsion, respectively. Natural detergents act as potent inhibitors of TuPL, and colipase restores the activity. When the lipase is inhibited by synthetic detergent, simultaneous addition of colipase and bile salts is required to restore the TuPL activity. The critical surface pressure of TuPL (pi(c)) = 20.9 mN m(-1)) is similar to the one of human PL (pi(c) = 18 mN m(-1)).
The results presented in this article indicate that despite the primitive character of the turtle, no significant difference has been observed between TuPL and known mammalian PLs. However, partial proteolysis of TuPL with chymotrypsin shows the absence of the 14-kDa fragment identified as the C-terminal domain in the case of many classic PLs.
本研究旨在从爬行动物海龟中纯化并鉴定经典胰脂肪酶（TuPL）。
从新鲜胰腺提取物中纯化脂肪酶，随后进行二乙氨基乙基纤维素色谱法、Sephacryl S - 200凝胶过滤法和Mono - Q琼脂糖色谱法。
海龟胰脂肪酶是一种丝氨酸酶，仅含有1个游离半胱氨酸。其活性在pH 8.2和37℃时最高。用三丁酸甘油酯和橄榄油乳液滴定法分别测得其比活性为10000 U/mg和5000 U/mg。天然洗涤剂是TuPL的强效抑制剂，而辅脂酶可恢复其活性。当脂肪酶被合成洗涤剂抑制时，需要同时添加辅脂酶和胆汁盐才能恢复TuPL活性。TuPL的临界表面压力（π(c)=20.9 mN m(-1)）与人胰脂肪酶的临界表面压力（π(c)=18 mN m(-1)）相似。
本文给出的结果表明，尽管海龟具有原始特征，但TuPL与已知的哺乳动物胰脂肪酶之间未观察到显著差异。然而，用胰凝乳蛋白酶对TuPL进行部分蛋白水解显示，在许多经典胰脂肪酶的情况下不存在被鉴定为C末端结构域的14 kDa片段。
