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Pancreatic Trypsin Cleaves Intestinal Alkaline Sphingomyelinase from Mucosa and Enhances the sphingomyelinase Activity.

Wu, Jun LU ; Liu, Fuli LU ; Nilsson, Åke LU and Duan, Rui-Dong LU (2004) In American Journal of Physiology: Gastrointestinal and Liver Physiology 287(5). p.967-973
Abstract
Sphingomyelin (SM) hydrolysis in the gut has implications in colonic tumorigenesis and cholesterol absorption. It is triggered by intestinal alkaline sphingomyelinase (Alk-SMase) that is present in the intestinal mucosa and content. The mechanism by which the enzyme is released into the lumen is not clear. We studied whether trypsin can dissociate Alk-SMase from the mucosa and affect its activity. During luminal perfusion of rat intestine, addition of trypsin to the buffer increased Alk-SMase activity in the perfusate output by about threefold. Treating COS-7 cells transfected with Alk-SMase cDNA with trypsin increased the SMase activity in the medium and reduced that in the cell lysate dose dependently. The appearance of Alk-SMase in the... (More)
Sphingomyelin (SM) hydrolysis in the gut has implications in colonic tumorigenesis and cholesterol absorption. It is triggered by intestinal alkaline sphingomyelinase (Alk-SMase) that is present in the intestinal mucosa and content. The mechanism by which the enzyme is released into the lumen is not clear. We studied whether trypsin can dissociate Alk-SMase from the mucosa and affect its activity. During luminal perfusion of rat intestine, addition of trypsin to the buffer increased Alk-SMase activity in the perfusate output by about threefold. Treating COS-7 cells transfected with Alk-SMase cDNA with trypsin increased the SMase activity in the medium and reduced that in the cell lysate dose dependently. The appearance of Alk-SMase in the perfusate and culture medium was confirmed by Western blot analysis. The effect of trypsin was blocked by trypsin inhibitor, and neither chymotrypsin nor elastase had a similar effect. We also expressed the full length and COOH-terminal truncated Alk-SMase in COS-7 cells and found that the activity of the full-length enzyme is mainly in the cells, whereas that of the truncated form is mainly in the medium. Both forms were active, but only the activity of the full-length Alk-SMase was enhanced by trypsin. By linking a poly-His tag to the constructed cDNA, we found that the first tryptic site Arg440 upstream of the signal anchor was attacked by trypsin. In conclusion, trypsin cleaves the Alk-SMase at the COOH terminal, releases it from mucosa, and meanwhile enhances its activity. The findings indicate a physiological role of trypsin in SM digestion. (Less)
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author
organization
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type
Contribution to journal
publication status
published
subject
in
American Journal of Physiology: Gastrointestinal and Liver Physiology
volume
287
issue
5
pages
967 - 973
publisher
American Physiological Society
external identifiers
  • wos:000224382600006
  • pmid:15205117
  • scopus:7044232133
ISSN
1522-1547
DOI
10.1152/ajpgi.00190.2004
language
English
LU publication?
yes
id
512075d1-f71c-48ab-ae0f-e9a033f50250 (old id 124082)
alternative location
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=15205117&dopt=Abstract
date added to LUP
2007-06-26 13:29:00
date last changed
2017-11-12 03:19:38
@article{512075d1-f71c-48ab-ae0f-e9a033f50250,
  abstract     = {Sphingomyelin (SM) hydrolysis in the gut has implications in colonic tumorigenesis and cholesterol absorption. It is triggered by intestinal alkaline sphingomyelinase (Alk-SMase) that is present in the intestinal mucosa and content. The mechanism by which the enzyme is released into the lumen is not clear. We studied whether trypsin can dissociate Alk-SMase from the mucosa and affect its activity. During luminal perfusion of rat intestine, addition of trypsin to the buffer increased Alk-SMase activity in the perfusate output by about threefold. Treating COS-7 cells transfected with Alk-SMase cDNA with trypsin increased the SMase activity in the medium and reduced that in the cell lysate dose dependently. The appearance of Alk-SMase in the perfusate and culture medium was confirmed by Western blot analysis. The effect of trypsin was blocked by trypsin inhibitor, and neither chymotrypsin nor elastase had a similar effect. We also expressed the full length and COOH-terminal truncated Alk-SMase in COS-7 cells and found that the activity of the full-length enzyme is mainly in the cells, whereas that of the truncated form is mainly in the medium. Both forms were active, but only the activity of the full-length Alk-SMase was enhanced by trypsin. By linking a poly-His tag to the constructed cDNA, we found that the first tryptic site Arg440 upstream of the signal anchor was attacked by trypsin. In conclusion, trypsin cleaves the Alk-SMase at the COOH terminal, releases it from mucosa, and meanwhile enhances its activity. The findings indicate a physiological role of trypsin in SM digestion.},
  author       = {Wu, Jun and Liu, Fuli and Nilsson, Åke and Duan, Rui-Dong},
  issn         = {1522-1547},
  language     = {eng},
  number       = {5},
  pages        = {967--973},
  publisher    = {American Physiological Society},
  series       = {American Journal of Physiology: Gastrointestinal and Liver Physiology},
  title        = {Pancreatic Trypsin Cleaves Intestinal Alkaline Sphingomyelinase from Mucosa and Enhances the sphingomyelinase Activity.},
  url          = {http://dx.doi.org/10.1152/ajpgi.00190.2004},
  volume       = {287},
  year         = {2004},
}