Quantitation of the endopeptidase activity generating γ-endorphin from β-endorphin in rat brain synaptic membranes by a radiometric assay

Lebouille, Jos L.M.; Burbach, J. Peter H.; Kloet, E. Ronald de

doi:10.1016/0003-2697(84)90417-2

Cited by 19 publications

(17 citation statements)

References 18 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The enzyme activity responsible for the cleavage has been term ed tentatively 'y-endorphin-generating endopeptidase' (yEGE) [2,21]. In order to quantitate this enzyme we have devised a l4C-labelled synthetic analogue of p-endorphin- (15)(16)(17)(18)(19) as a substrate [21].…”

mentioning

confidence: 99%

Leu-Phe Cleaving Endopeptidase Activity, γ-Endorphin, and β-Endorphin in the Rat Pituitary Gland and Brain

Lebouille¹,

Burbach²,

Kloet³

et al. 1988

Neuroendocrinology

Self Cite

View full text Add to dashboard Cite

The influence of adrenalectomy and corticosterone substitution was investigated on Leu-Phe cleaving endopeptidase activity and on the levels of γ-endorphin and β-endorphin in the pituitary gland and the brain. The enzyme activity was quantitated by a specific radiometric assay based on the cleavage of the Leu¹⁷-Phe¹⁸ bond in a NH₂ and COOH-terminally protected synthetic substrate wich was analogous to β-endorphin-(15–19). This activity may mimick the formation of γ-endorphin. β-Endorphin and γ-endorphin were measured by specific radioimmunoassays. After 14 days of adrenalectomy enzyme activity had increased in anterior (15%) and neurointermediate lobes of the pituitary gland (30%), hypothalamus (25%), and liver (15%). This increase was prevented when the adrenalectomized animals were subjected to chronic corticosterone substitution by subcutaneous implantation of a pellet of 100 mg. Extirpation of only the adrenal medulla did not affect the Leu-Phe cleaving activity. Enzyme activity in the septum, hippocampus, and cerebellum had not changed after adrenalectomy. Determination of immunoreactive levels of γ- and β-endorphins showed that in the anterior pituitary gland γ- and β-endorphins had increased by 275 and 300%, respectively, 14 days after adrenalectomy. No significant changes were observed in endorphin levels of the intermediate lobe of the pituitary gland, hypothalamus, hippocampus, and septum. The results indicate that Leu-Phe cleaving endopeptidase activity is sensitive to glucocorticoids in tissues containing proopiomelanocortin-producing cells, i.e., anterior and neurointermediate pituitary gland and the hypothalamus. In the anterior pituitary gland it is correlated with the levels of γ- and β-endorphins.

show abstract

mentioning

confidence: 99%

Leu-Phe Cleaving Endopeptidase Activity, γ-Endorphin, and β-Endorphin in the Rat Pituitary Gland and Brain

Lebouille¹,

Burbach²,

Kloet³

et al. 1988

Neuroendocrinology

Self Cite

View full text Add to dashboard Cite

show abstract

“…The substrate was N-and C-terminally protected against exopeptidases. The substrate was r elabeled by reductive methylation of the €-NH 2 group of the Lys residue with [ 14 C]-formaldehyde and anhydrous sodium cyanoborohydride as reductive agent (24). With two 14 C-labeled-CH 3 groups incorporated, the specific radioactivity of the 14 C-labeled substrate was 106 mCi/mmol.…”

Section: Assay Of Yege Activitymentioning

confidence: 99%

“…The assay of 7EGE activity was performed as previously dexcribed (24,25). Briefly, a synthetic substrate AC-Val-ThrLeu -Phe -Lys([ 14 C] CH 3 ) 2 -NHCH 3 (Ac -VTLFK([ 14 C]CH 3 ) 2 -NHCH 3 ), a derivative of /3-endorphin- (15)(16)(17)(18)(19), was used.…”

Section: Assay Of Yege Activitymentioning

confidence: 99%

“…7-Endorphin is generated from 0-endorphin by a single cleavage of the Leu 17 -Phe 18 bond by a distinct endopeptidase (21)(22)(23)(24). We termed this enzyme activity 7-endorphin-generating endopeptidase (7EGE) activity (24,25).…”

mentioning

confidence: 99%

“…We termed this enzyme activity 7-endorphin-generating endopeptidase (7EGE) activity (24,25). To investigate the biological significance of 7EGE activity in the metabolism of j8-endorphin, an assay for 7EGE activity was developed.…”

mentioning

confidence: 99%

See 2 more Smart Citations

γ-Endorphin-Generating Endopeptidase: Distribution in Body Tissues and Cellular Localization in Rat Testis*

et al. 1986

View full text Add to dashboard Cite

The Leu17-Phe18 bond of beta-endorphin is cleaved by a specific endopeptidase that generates the biologically active peptide gamma-endorphin. gamma-Endorphin-generating endopeptidase (gamma EGE) activity was determined by a radiometric assay, using as substrate a radioactively labeled, N- and C-terminally protected pentapeptide: Ac-Val-Thr-Leu-Lys( [14C]CH3)2-NHCH3, a derivative of beta-endorphin-(15-19). Here we report the tissue distribution of gamma EGE activity and its cellular localization in the testis. gamma EGE activity was present in the cytosolic fraction of most tissues. Highest specific activity occurred in the testis, ovary, and the uterus (10-16 nmol X mg protein-1 X h-1). In testis highest specific gamma EGE activity was found in the tubules (42 nmol X mg protein-1 X h-1) and lowest in Leydig cells (8 nmol X mg protein-1 X h-1). Further fractionation of the tubules showed that the germinal cell fraction had a higher specific activity (24 nmol X mg protein-1 X h-1) than the Sertoli cell fraction (8 nmol X mg protein-1 X h-1). In testis depleted of the germinal cells by prenatal irradiation of the rat or hypophysectomy, specific activity of gamma EGE activity decreased 50-fold and 4-fold, respectively. In testis depleted of Leydig cells by treatment of rats with ethane dimethyl sulfonate, specific gamma EGE activity did not decrease. Adrenalectomy had no effect on the enzyme activity. The results suggest that the germinal cells are sites of processing of beta-endorphin into alpha- and gamma-endorphins. It is concluded that 1) gamma EGE activity is widely distributed in tissues; 2) highest gamma EGE activity is located in reproductive tissues; and 3) in the testis gamma EGE activity is mainly associated in the germinal cells.

show abstract

Dopaminergic Agents Have Differential Effects on Both Content and Processing of Alpha-N-Acetylated Endorphin in the Ovine Pituitary Intermediate Lobe

Smith¹,

Wallace²,

Clarke³

et al. 1989

Neuroendocrine Perspectives

View full text Add to dashboard Cite

Quantitation of the endopeptidase activity generating γ-endorphin from β-endorphin in rat brain synaptic membranes by a radiometric assay

Cited by 19 publications

References 18 publications

Leu-Phe Cleaving Endopeptidase Activity, γ-Endorphin, and β-Endorphin in the Rat Pituitary Gland and Brain

Leu-Phe Cleaving Endopeptidase Activity, γ-Endorphin, and β-Endorphin in the Rat Pituitary Gland and Brain

γ-Endorphin-Generating Endopeptidase: Distribution in Body Tissues and Cellular Localization in Rat Testis*

Dopaminergic Agents Have Differential Effects on Both Content and Processing of Alpha-N-Acetylated Endorphin in the Ovine Pituitary Intermediate Lobe

Contact Info

Product

Resources

About