Demonstration of γ-glutamyl transpeptidase activity in human jejunal mucosa

Greenberg, Elaine; Wollaeger, Eric E.; Fleisher, Gerard A.; Engström, Gunnar

doi:10.1016/0009-8981(67)90272-0

Cited by 49 publications

(15 citation statements)

References 21 publications

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“…The present findings, which indicate that y-glutamyl transpeptidase is a villus-specific enzyme, are in accord with earlier histochemical studies in which the products of this enzyme's activity were found to be localized in the brush border regions of the human (27), rabbit (28), and guinea pig (29) small intestine. In contrast to such localization of y-glu- in accord with the view that the transpeptidase catalyzes the quantitatively major pathway of glutathione breakdown, and strongly suggests that such utilization of glutathione is connected with a significant physiological activity of the villus tip cells.…”

Section: Discussionsupporting

confidence: 92%

Glutathione and gamma-glutamyl cycle enzymes in crypt and villus tip cells of rat jejunal mucosa.

Cornell¹,

Meister²

1976

Proc. Natl. Acad. Sci. U.S.A.

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Villus tip cells and crypt cells of rat jejunal mucosa were separated bythe planing procedure of Imondi et a]. and were studied with respect to their activities of the enzymes of the 'y-glutamyl cycle and glutathione content.The villus tip cells exhibit much higher y-glutamyl transpeptidase activities than do the crypt cells; thus, y-glutamyl transpeptidase appears to be a villus-specific enzyme. y-Glutamyl cyclotransferase and the enzymes required for glutathione synthesis are not specifically localized to In the present work, the micrometer planing technique devised by Imondi et al. (7) was used to separate the epithelial cells of rat jejunum. The intracellular concentration of glutathione and the activities of enzymes involved in glutathione metabolism were examined; two of these ('y-glutamyl cysteine synthetase, glutathione synthetase) catalyze the synthesis of glutathione from its constituent amino acids, and two (y-glutamyl transpeptidase, y-glutamyl cyclotransferase) are involved in the major degradative pathway of glutathione metabolism (17). The uptake of amino acids by the isolated crypt and villus cells was also studied. Methods. Male Sprague-Dawley rats (250-300 g) were housed in stainless steel cages with wire mesh bottoms; the animals were fed ad lib. on standard laboratory rat chow. Groups of rats were fasted for 24 or 48 hr; some were fasted for 24 hr, and then fed ad lib. with cane sugar cubes. Other rats were fasted for 24 hours then fed ad lib. a protein mixture prepared by mixing 8.0 g of gelatin (Knox) and 10 g of casamino acids (Difco) with 400 ml of water.The animals were sacrificed by decapitation and their small intestines were excised. The duodenum was removed and the first 18 cm of the jejunum was cut into three sections, each of which was mounted on the tissue planing device and planed at 125 ,um depths according to the method of Imondi et al. (7). The fractions were quick-frozen in petri dishes placed on dry ice. Five fractions were obtained from the animals fed ad lib. For the enzyme studies the cells were thawed and homogenized in five volumes (weight/volume) of 10 mM MgCl2, 10 mM Tris-HCl (pH 7.6). The homogenate was layered on 18% sucrose containing 10 mM MgCl2 and 10 mM Tris-HCl (pH 7.6) and centrifuged at 100,000 X g to sediment y-glutamyl transpeptidase. y-Glutamyl transpeptidase was assayed after diluting the pellet to an appropriate volume with 0.1 M Tris-HCl (pH 8.0). The other enzymes were assayed in the supernatant collected at the top of the sucrose layer. Protein was determined by the method of Lowry (20) using bovine serum albumin as standard. Thymidine kinase was assayed by the method of Klemperer and Haynes (21) except that thymidine and thymidine 5'-phosphate were separated by ascending paper chromatography in tertiary butanol:methylethylketone:water:formic acid (44:44:11:0.26; vol/vol) (22). -y-Glutamyl transpeptidase was assayed with L-y-glutamyl-p-nitroanilide and glycylglycine 420

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Section: Discussionsupporting

confidence: 92%

Glutathione and gamma-glutamyl cycle enzymes in crypt and villus tip cells of rat jejunal mucosa.

Cornell¹,

Meister²

1976

Proc. Natl. Acad. Sci. U.S.A.

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“…GGT has been found at highest activities in plasma membranes of epithelia specialised in absorptive functions, such as the proximal nephron and the small intestine, where the enzyme seems to be localised in the brush border membrane . In contrast GGT is virtually absent from colonic epithelium [6].…”

Section: Introductionmentioning

confidence: 89%

γ‐glutamyl transferase activity in the pig proximal colon during early postnatal development

Sepúlveda

Burton

1982

FEBS Letters

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“…I t catalyzes both the hydrolysis of y glutamyl-containing peptide bonds and the transfer of y glutamyl groups from donor peptides to suitable acceptors, i. e. transpeptidation. GGTP activity has been found in many tissues [3, 1 1, 13,23,27] of small laboratory animals [ll], hog [26], cow [20], sea lion [2], and man 13,11,13,23,271. Techniques originally developed for estimating GGTP activity involved quantitating the disappearance of glutathione and the appearance of the products of glutathione cleavage [4,8,28].…”

Section: Introductionmentioning

confidence: 99%

“…I n the small intestine it is localized to the mucosal brush border membrane. This localization suggests a possible role in synthesis, degradation, and/or trans- Gestat~onal age (days) Post natal age (days) 13 9 24…”

mentioning

confidence: 98%

“…The biochemical and physiological significance of GGTP is uncertain. Some authors have suggested a role in the transport of amino acids across cell membranes [13], in the regulation of tissue levels of glutathione [16], and in the peptide-nitrogen storage [16, 301. This enzyme is present in many organs and specifically in various portions of the gastrointestinal tract. I n the small intestine it is localized to the mucosal brush border membrane.…”

mentioning

confidence: 99%

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Gamma Glutamyl Transpeptidase: Measurement and Development in Guinea Pig Small Intestine

et al. 1969

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ExtractA modification ofa previous method for the assay ofy glutamyI transpeptidase (GGTP) was developed.Substrate solubility difficulties alluded to by other investigators were avoided by employing heating and solubilization of the chromogenic substrate y glutamyl-P-naphthylamide in a medium of carbonate 0.05 M and Tris buffer 0.1 M a t p H 9.5. The kinetics and conditions for such an assay are described. Whole intestinal homogenates of adult male guinea pigs were used as the source of the enzyme.The developmental pattern of this enzyme was determined in fetuses at 55 and 63 days of gestation and at varying times from 1 to 90 days of age. A total of 69 animals was assayed. The general pattern was that ofhigh specific activity during prenatal life, with a rapid decline during the neonatal period (3-24 days of age) and a slight increase after 55 days of age.Other guinea pig organs were studied. Liver and kidney were found to contain enzyme activity greater than that of the intestine. Subcellular fractionation of intestinal mucosa using ultracentrifugation revealed a twenty-fold enrichment of activity in jejunal brush border membrane, compared with whole jejunal homogenates when expressed as specific enzyme activity per mg of protein. The stability characteristics of GGTP disclosed no loss of specific activity when stored a t -28' for 50 days.This simple enzyme assay, stability of the enzyme when frozen, subcellular distribution in the intestinal brush border membrane, and an unusual developmental pattern made this enzyme a useful adjunct to the study of intestinal protein metabolism. SpeculationThe unique ability of GGTP to hydrolyze y glutamic acid-peptide bonds and the location of the enzyme in the intestinal brush border suggest a role for this enzyme in the metabolism of an unusual group of physiologically important peptides such as glutathione, folic acid, and gluten-gliadin. The significance of these y-bonded compounds may now be approached in order to investigate the role of this enzyme in gluten enteropathy and related disorders.

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Demonstration of γ-glutamyl transpeptidase activity in human jejunal mucosa

Cited by 49 publications

References 21 publications

Glutathione and gamma-glutamyl cycle enzymes in crypt and villus tip cells of rat jejunal mucosa.

Glutathione and gamma-glutamyl cycle enzymes in crypt and villus tip cells of rat jejunal mucosa.

γ‐glutamyl transferase activity in the pig proximal colon during early postnatal development

Gamma Glutamyl Transpeptidase: Measurement and Development in Guinea Pig Small Intestine

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