Dudeja P K, Brasitus T A
Department of Medicine, University of Illinois, Chicago.
Gastroenterology. 1993 Aug;105(2):357-66. doi: 10.1016/0016-5085(93)90708-k.
To date, the potential effects of free radicals on the activities of intestinal brush-border membrane enzymes have received little attention. Therefore, we conducted a series of experiments to examine the effects of free radicals on various enzymatic activities of rat small intestinal brush-border membranes.
An in vitro Fe2+/ascorbate oxygen-radical generating system and rat small intestinal brush-border membranes were used for this purpose. The rate of lipid peroxidation was used as an index of free radical damage. In addition, fluorescence polarization, fatty acid analyses, membrane delipidation techniques, and studies with antioxidants and metal cofactors were used.
Increased free radical formation was associated with the inhibiton of alkaline phosphatase activity, with no change in the activities of sucrase, maltase, leucine aminopeptidase, and gamma-glutamyl transpeptidase. Generation of free radicals also markedly decreased membrane lipid fluidity and altered fatty acid composition. Catalase, reduced glutathione, or alpha-tocopherol treatment reduced lipid peroxidation as well as inactivation of enzyme activity. The use of artificial fluidizers, Arrhenius plots, and membrane delipidation studies failed to show a relationship between the inactivation of this enzyme and changes in membrane lipid composition or fluidity.
The inactivation of alkaline phosphatase by free radicals appears to involve its direct oxidation.
迄今为止，自由基对肠刷状缘膜酶活性的潜在影响鲜受关注。因此，我们开展了一系列实验来研究自由基对大鼠小肠刷状缘膜各种酶活性的影响。
为此使用了体外Fe2+/抗坏血酸氧自由基生成系统和大鼠小肠刷状缘膜。脂质过氧化速率用作自由基损伤的指标。此外，还使用了荧光偏振、脂肪酸分析、膜脱脂技术以及抗氧化剂和金属辅因子研究。
自由基形成增加与碱性磷酸酶活性受到抑制相关，而蔗糖酶、麦芽糖酶、亮氨酸氨肽酶和γ-谷氨酰转肽酶的活性没有变化。自由基的产生还显著降低了膜脂质流动性并改变了脂肪酸组成。过氧化氢酶、还原型谷胱甘肽或α-生育酚处理可减少脂质过氧化以及酶活性的失活。使用人工流化剂、阿累尼乌斯图和膜脱脂研究未能表明该酶的失活与膜脂质组成或流动性变化之间存在关联。
自由基导致碱性磷酸酶失活似乎涉及其直接氧化。
