Evidence for specific ceramidase present in the intestinal contents of rats and humans
Evidence for specific ceramidase present in the intestinal contents of rats and humans
复制标题
DOI：
10.1007/s11745-001-0788-3
复制
发表时间：
2001-08-01
期刊：
影响因子：
1.9
通讯作者：
Nilsson, Å
中科院分区：
文献类型：
--
作者：
Duan, RD;Cheng, YJ;Nilsson, Å
先前在肠道内容物中发现了由胆汁盐刺激的中性神经酰胺酶活性。最近，胆盐刺激脂肪酶（BSSL）被发现具有神经酰胺酶活性。目前尚不清楚先前发现的神经酰胺酶活性是否归因于 BSSL。为了解决这个问题，我们比较了神经酰胺酶和胰腺 BSSL 的高季氨乙基 (HQ) 阴离子交换色谱的行为、分布、稳定性以及对脂肪酶抑制剂的反应。来自人和大鼠整个小肠内容物的蛋白质用丙酮沉淀并溶解在 20 mM Tris 缓冲液 pH 8.2 中。这些蛋白质具有中性神经酰胺酶活性，但不具有针对乙酸对硝基苯酯的 BSSL 活性。当蛋白质进行 HQ 层析时，鉴定出神经酰胺酶活性的两个峰，分别具有酸性和中性 pH 最佳值。它们均不具有针对乙酸对硝基苯酯的 BSSL 活性。使用 BSSL 抗血清的蛋白质印迹未能鉴定具有高中性神经酰胺酶活性的级分中的 BSSL 蛋白。在大鼠肠道中，胰腺BSSL活性在十二指肠中较高，在小肠中迅速下降，而中性神经酰胺酶活性在十二指肠中较低，直到小肠远端一直保持较高水平。此外，脂肪酶抑制剂奥利司他消除了人BSSL对乙酸对硝基苯酯的活性，并略微降低了其对神经酰胺的活性，但对HQ色谱分离的神经酰胺酶活性没有抑制作用。总之，我们提供了肠道内容物中存在除胰腺 BSSL 之外的特定神经酰胺酶的证据。该酶可能在膳食鞘脂的消化中发挥重要作用。
A neutral ceramidase activity stimulated by bile salt was previously identified in the intestinal content. Recently, bile salt stimulated lipase (BSSL) was found to have ceramidase activity. It is unknown whether the ceramidase activity previously found is attributable to BSSL. To address this question, we compared the behaviors of high quaternary aminoethyl (HQ) anion exchange chromatography, the distributions, the stability, and the responses to lipase inhibitor between ceramidase and pancreatic BSSL. The proteins from whole small intestinal contents of humans and rats were precipitated by acetone and dissolved in 20 mM Tris buffer pH 8.2. These proteins had neutral ceramidase activity but not BSSL activity against p-nitrophenyl acetate. When the proteins were subject to HQ chromatography, two peaks of ceramidase activity were identified, which had acid and neutral pH optima, respectively. Neither of them had BSSL activity against p-nitrophenyl acetate. Western blot using BSSL antiserum failed to identify BSSL protein in the fractions with high neutral ceramidase activity. In rat intestinal tract, pancreatic BSSL activity was high in the duodenum and declined rapidly in the small intestine, whereas neutral ceramidase activity was low in the duodenum and maintained a high level until the distal part of the small intestine. In addition, orlistat, the inhibitor of lipase, abolished human BSSL activity against p-nitrophenyl acetate and slightly reduced its activity against ceramide but had no inhibitory effect on ceramidase activity isolated by HQ chromatography. In conclusion, we provide the evidence for a specific ceramidase other than pancreatic BSSL present in the intestinal content. The enzyme may play important roles in digestion of dietary sphingolipids.