Pancreatic Trypsin Cleaves Intestinal Alkaline Sphingomyelinase from Mucosa and Enhances the sphingomyelinase Activity.
(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)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/124082
- author
- Wu, Jun LU ; Liu, Fuli LU ; Nilsson, Åke LU and Duan, Rui-Dong LU
- organization
- publishing date
- 2004
- 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
- 2016-04-01 11:41:51
- date last changed
- 2024-01-07 17:03:14
@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}}, doi = {{10.1152/ajpgi.00190.2004}}, volume = {{287}}, year = {{2004}}, }