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Purification and characterization of a luminal cholecystokinin-releasing factor from rat intestinal secretion.

Publication ,  Journal Article
Spannagel, AW; Green, GM; Guan, D; Liddle, RA; Faull, K; Reeve, JR
Published in: Proc Natl Acad Sci U S A
April 30, 1996

Cholecystokinin (CCK) secretion in rats and humans is inhibited by pancreatic proteases and bile acids in the intestine. It has been hypothesized that the inhibition of CCK release caused by pancreatic proteases is due to proteolytic inactivation of a CCK-releasing peptide present in intestinal secretion. To purify the putative luminal CCK-releasing factor (LCRF), intestinal secretions were collected by perfusing a modified Thiry-Vella fistula of jejunum in conscious rats. From these secretions, the peptide was concentrated by ultrafiltration followed by low-pressure reverse-phase chromatography and purified by reverse-phase high-pressure liquid chromatography. Purity was confirmed by high-performance capillary electrophoresis. Fractions were assayed for CCK-releasing activity by their ability to stimulate pancreatic protein secretion when infused into the proximal small intestine of conscious rats. Partially purified fractions strongly stimulated both pancreatic secretion and CCK release while CCK receptor blockade abolished the pancreatic response. Amino acid analysis and mass spectral analysis showed that the purified peptide is composed of 70-75 amino acid residues and has a mass of 8136 Da. Microsequence analysis of LCRF yielded an amino acid sequence for 41 residues as follows: STFWAYQPDGDNDPTDYQKYEHTSSPSQLLAPGDYPCVIEV. When infused intraduodenally, the purified peptide stimulated pancreatic protein and fluid secretion in a dose-related manner in conscious rats and significantly elevated plasma CCK levels. Immunoaffinity chromatography using antisera raised to synthetic LCRF-(1-6) abolished the CCK releasing activity of intestinal secretions. These studies demonstrate, to our knowledge, the first chemical characterization of a luminally secreted enteric peptide functioning as an intraluminal regulator of intestinal hormone release.

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Published In

Proc Natl Acad Sci U S A

DOI

ISSN

0027-8424

Publication Date

April 30, 1996

Volume

93

Issue

9

Start / End Page

4415 / 4420

Location

United States

Related Subject Headings

  • Reproducibility of Results
  • Rats, Wistar
  • Rats
  • Proteins
  • Pancreatic Juice
  • Molecular Sequence Data
  • Mass Spectrometry
  • Male
  • Jejunum
  • Intestinal Mucosa
 

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Spannagel, A. W., Green, G. M., Guan, D., Liddle, R. A., Faull, K., & Reeve, J. R. (1996). Purification and characterization of a luminal cholecystokinin-releasing factor from rat intestinal secretion. Proc Natl Acad Sci U S A, 93(9), 4415–4420. https://doi.org/10.1073/pnas.93.9.4415
Spannagel, A. W., G. M. Green, D. Guan, R. A. Liddle, K. Faull, and J. R. Reeve. “Purification and characterization of a luminal cholecystokinin-releasing factor from rat intestinal secretion.Proc Natl Acad Sci U S A 93, no. 9 (April 30, 1996): 4415–20. https://doi.org/10.1073/pnas.93.9.4415.
Spannagel AW, Green GM, Guan D, Liddle RA, Faull K, Reeve JR. Purification and characterization of a luminal cholecystokinin-releasing factor from rat intestinal secretion. Proc Natl Acad Sci U S A. 1996 Apr 30;93(9):4415–20.
Spannagel, A. W., et al. “Purification and characterization of a luminal cholecystokinin-releasing factor from rat intestinal secretion.Proc Natl Acad Sci U S A, vol. 93, no. 9, Apr. 1996, pp. 4415–20. Pubmed, doi:10.1073/pnas.93.9.4415.
Spannagel AW, Green GM, Guan D, Liddle RA, Faull K, Reeve JR. Purification and characterization of a luminal cholecystokinin-releasing factor from rat intestinal secretion. Proc Natl Acad Sci U S A. 1996 Apr 30;93(9):4415–4420.
Journal cover image

Published In

Proc Natl Acad Sci U S A

DOI

ISSN

0027-8424

Publication Date

April 30, 1996

Volume

93

Issue

9

Start / End Page

4415 / 4420

Location

United States

Related Subject Headings

  • Reproducibility of Results
  • Rats, Wistar
  • Rats
  • Proteins
  • Pancreatic Juice
  • Molecular Sequence Data
  • Mass Spectrometry
  • Male
  • Jejunum
  • Intestinal Mucosa