Ville Eric, Carrière Frédéric, Renou Christophe, Laugier René
La Timone Hospital, Gastroenterology Department, Marseille, France.
Digestion. 2002;65(2):73-81. doi: 10.1159/000057708.
BACKGROUND/AIMS: The aim of this study was to exactly determine the pH stability of human gastric lipase (HGL) and to investigate the mechanism underlying the inactivation of HGL which occurs in gastric juice.
Samples of human gastric juice and purified HGL were incubated at various pH values ranging from 0.5 to 8.0, and the residual HGL activity was measured as a function of time using the pHstat technique. Samples of purified HGL were also incubated in the presence of human pepsin. Electrophoresis and Western blot analysis were performed on all the samples in which HGL was inactivated.
HGL was found to be stable in gastric juice at pH values ranging from 2.0 to 7.0, especially between pH 3.0 and 5.0 (half-inactivation time >24 h). HGL activity decreased rapidly below pH 2.0 and above pH 7.0. The inactivation half times were only 43 +/- 9 and 24 +/- 18 min at pH 1 and pH 8, respectively. The pH stability of purified HGL was much lower than that of HGL in gastric juice. Acid or alkaline inactivation of HGL could occur without any prior proteolytic degradation, and this inactivation was irreversible. However, proteolytic degradation of HGL by pepsin also occurred at very low pH values, probably because the acid-denatured HGL is more sensitive to proteolytic cleavage by pepsin. An ex vivo study of HGL activity in several gastric juice samples showed that the HGL activity decreased with the pH of the sample, in both basal and pentagastrin-stimulated gastric juice.
Although HGL is not as stable as it was previously thought to be under acidic conditions, it is nevertheless the most stable acid lipase and constitutes a good candidate tool for enzyme substitution therapy.
背景/目的：本研究旨在准确测定人胃脂肪酶（HGL）的pH稳定性，并探究胃液中发生的HGL失活机制。
将人胃液和纯化的HGL样品在pH值为0.5至8.0的不同条件下孵育，使用pH计技术测定残余HGL活性随时间的变化。纯化的HGL样品也在人胃蛋白酶存在的情况下孵育。对所有HGL失活的样品进行电泳和蛋白质印迹分析。
发现HGL在pH值为2.0至7.0的胃液中稳定，特别是在pH 3.0至5.0之间（半失活时间>24小时）。在pH值低于2.0和高于7.0时，HGL活性迅速下降。在pH 1和pH 8时，失活半衰期分别仅为43±9分钟和24±18分钟。纯化的HGL的pH稳定性远低于胃液中的HGL。HGL的酸或碱失活可在没有任何先前蛋白水解降解的情况下发生，并且这种失活是不可逆的。然而，胃蛋白酶对HGL的蛋白水解降解也在非常低的pH值下发生，可能是因为酸变性的HGL对胃蛋白酶的蛋白水解切割更敏感。对几个胃液样品中HGL活性的体外研究表明，在基础胃液和五肽胃泌素刺激的胃液中，HGL活性均随样品pH值的降低而降低。
尽管HGL在酸性条件下不如先前认为的稳定，但它仍然是最稳定的酸性脂肪酶，是酶替代疗法的良好候选工具。
