Evidence for specific ceramidase present in the intestinal contents of rats and humans.
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.
Duan
R D
RD
Department of Cell Biology B, Biomedical Center, University of Lund, Sweden. Rui-dong.duan@med.lu.se
Cheng
Y
Y
Yang
L
L
Ohlsson
L
L
Nilsson
A
A
D000581:Amidohydrolases / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D055572:Ceramidases; D002852:Chromatography, Ion Exchange / Q000379:methods; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007421:Intestine, Small / Q000378:metabolism; D007422:Intestines / Q000201:enzymology*; D007783:Lactones / Q000494:pharmacology; D055576:Neutral Ceramidase; D000077403:Orlistat; D051381:Rats; D017207:Rats, Sprague-Dawley; D002787:Sterol Esterase / Q000037:antagonists & inhibitors / Q000378:metabolism
D004791:Enzyme Inhibitors; D007783:Lactones; D000077403:Orlistat; C054784:bile salt-stimulated lipase; D002787:Sterol Esterase; D000581:Amidohydrolases; C526744:ASAH2 protein, human; C526748:Asah2 protein, rat; D055572:Ceramidases; D055576:Neutral Ceramidase