Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01189 (beta-endorphin)
 21,003 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Binding of human beta-endorphin to mouse brain membrane preparations has been characterized using the tritiated hormone as primary ligand. The binding was shown to be time and temperature dependent. The dissociation constant of the saturable binding was determined to be 0.5 nM. Both [Met5]enkephalin and naloxone were found to compete with tritiated beta-endorphin for the binding with a potency of 6.3 and 6.5% of the unlabeled hormone, respectively. Phospholipase A2 isolated from Formosan cobra venom was shown to be a potent inhibitor of the binding with a 50% inhibition concentration of 26 nM. Although Na+, K+, Ca2+, Mg2+ were found individually to exert a profound inhibition on the binding, the combined salt solution (Tyrode) did not abolish the specific binding of tritiated beta-endorphin to the mouse brain membrane preparations.
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PMID:beta-Endorphin: characteristics of binding sites in the mouse brain. 630 56

This study was designed to assess practically the suitability of different C18 reversed-phase radially compressed polythene cartridges (Radial-Pak, Waters Assoc.) in two types of radial-compression systems, for the separation and analysis of various neuropeptides at both high (less than 5 micrograms) and low (greater than 100 pg) levels in biological extracts and to compare them with well established techniques using stainless-steel columns. A solvent system fully compatible with both radially compressed and steel columns is described. The completely volatile mobile phase (acetonitrile gradient containing trifluoroacetic acid) allows ultraviolet detection below 215 nm, gives good resolution and is readily compatible with the further radioimmunoassay and bioassay of collected fractions. The efficiency of radially compressed 5 and 10 microns "capped" and "non-capped" C18 silica supports and slurry-packed steel columns has been assessed by: (1) separation and recovery of a complex standard mixture of neuropeptides; (2) separation and subsequent identification of degradation products formed during the incubation of neurotensin with rat cortical synaptosomes; (3) analysis of alpha-melanotropin and corticotropin-(18-39) in tissue culture media containing varying amounts of foetal calf serum; and (4) characterization of corticotropin-like immunoreactivity in human cerebrospinal fluid. The Z-module fitted with the capped 10-microns irregular C18 silica cartridge gave better resolution than with the mu Bondapak steel column but the selective retention was similar. The back-pressures in the Z-module are much reduced (approximately 13 bar at 1 ml/min); therefore, flow-rates may be increased and analysis times greatly reduced. In order to obtain good resolution with the RCM-100 module which uses a non-capped stationary phase, a salt must be added (e.g. 15 mM sodium chloride) to the mobile phase to reduce polar interactions between the peptide and the free silanol groups on the stationary phase. This makes the solvent non-volatile and therefore less useful.
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PMID:High-performance liquid chromatography of neuropeptides using radially compressed polythene cartridges. 632 86

Secretory granules (SGs) from rat intermediate lobes (IL) were isolated in a highly purified form by differential centrifugation, followed by sucrose density gradient centrifugation. The purified IL-SGs were lysed by freezing and thawing. The granule lysate was then centrifuged to generate membrane and soluble fractions. Proopiocortin -converting enzyme (PCE) activity was assayed by incubation of [3H]arginine- or [3H] phenylalanine-labeled toad proopiocortin with the total granule lysate, the membrane, or the soluble fraction at pH 5.0. The processed products were identified by immunoprecipitation with ACTH and beta-endorphin antisera, followed by acid-urea-gel electrophoresis. The PCE activity in rat IL-SG lysate cleaved proopiocortin to 21,000 mol wt ACTH, 21,000 mol wt ACTH/lipotropin (LPH), 13,000 mol wt ACTH, beta LPH, beta-endorphin-like peptides, and alpha MSH-like peptides, similar to those synthesized by the toad intermediate lobe in situ. Treatment of the PCE cleavage products with carboxypeptidase B resulted in the liberation of free arginine. This observation together with the nature of the products formed suggest that the PCE activity cleaved at pairs of basic residues of proopiocortin , yielding one or more products that terminated with an arginine or an arginine-lysine. PCE activity was found in membrane and soluble granule fractions, and both activities were inhibited by leupeptin, p-chloromercuribenzoate, dithiodipyridine, and pepstatin A, but not by chloroquine or N-alpha-p-tosyl-L-lysine-chloromethylketone HCl. Diisopropyl fluorophosphate and other thiol protease reagents (p-chloromercuriphenyl sulfonic acid, iodoacetic acid, and HgCl2) had a small inhibitory effect. The products formed by PCE activities in the membrane and soluble fractions were similar to those cleaved by the total granule lysate. The membrane fraction primarily cleaved proopiocortin between ACTH and beta LPH to form 21,000 (21 K) mol wt ACTH and beta-LPH, similar to the first processing step in the IL in situ. The soluble fraction, however, showed a greater tendency to cleave proopiocortin between the 16 K N-terminal glycopeptide and ACTH, to yield twice as much 21 K ACTH/LPH product as the membrane fraction. The membrane-associated PCE activity was found to be easily solubilized by extraction with high salt (1 M NaCl), suggesting that it is not an integral granule membrane protein.
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PMID:In vitro processing of proopiocortin by membrane-associated and soluble converting enzyme activities from rat intermediate lobe secretory granules. 632 33

Strips of rabbit pulmonary artery and aorta were incubated with [3H]noradrenaline and subsequently superfused. Tritium overflow from strips superfused with physiological salt solution was stimulated either electrically (usually at a frequency of 2 Hz) or by tyramine 1 mumol/l and overflow from strips superfused with Ca2+-free solution containing K+ 54.7 mmol/l was stimulated by introduction of Ca2+ 1.6 mmol/l. In most of the experiments (stimulation by electrical impulses or CaCl2) neuronal and extraneuronal uptake and beta-adrenoceptors were blocked by cocaine, corticosterone and propranolol, respectively. The electrically evoked overflow of 3H-labelled substances from pulmonary artery and aorta was increased by adrenocorticotropic hormone. In the pulmonary artery, the adrenocorticotropic hormone--induced increase in impulse--evoked overflow (and contraction) was the more pronounced, the lower the frequency of stimulation (6, 2 and 0.66 Hz: 360 impulses). The electrically evoked overflow of 3H-labelled substances was also increased by alpha-melanocyte-stimulating hormone and porcine adrenocorticotropic hormone, but was not affected by adrenocorticotropic hormone. Adrenocorticotropic hormone also facilitated the Ca2+-evoked overflow of 3H-labelled substances promoted by high K+, but it did not affect the Ca2+-independent tyramine-evoked overflow. Adrenocorticotropic hormone did not alter the percentages of [3H]noradrenaline and 3H-labelled metabolites contained in electrically or tyramine-evoked overflow of 3H-labelled substances. In conclusion, adrenocorticotropic hormone and fragments of adrenocorticotropic hormone cause an increase in stimulation-evoked, Ca2+-dependent noradrenaline release from postganglionic sympathetic nerve fibres, probably by activating presynaptic receptors for adrenocorticotropic hormone.
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PMID:Facilitatory effect of adrenocorticotropic hormone and related peptides on Ca2+-dependent noradrenaline release from sympathetic nerves. 633 Jun 7

The neurointermediary lobes from 190 rat pituitaries were homogenized in an acidic medium which inhibits peptidase activity and maximizes the solubilization of undamaged peptides. Octadecylsilyl-silica (ODS-silica) was used to extract the supernatant of the tissue homogenate. The ODS-silica eluate, now largely protein and salt free, was subjected to reversed-phase high-performance liquid chromatography (HPLC) employing 0.1% trifluoroacetic as counter ion. The column eluates were monitored for beta-endorphin immunoreactivity. Five immunoreactive components were observed. The most hydrophobic of these was repurified on the same HPLC column using 0.13% heptafluorobutyric acid as counter ion. Characterization of the purified peptide by gel permeation HPLC, amino acid analysis, and tryptic fragmentation indicated that it corresponded in structure to alpha-N-acetyl-beta-endorphin1-26. Amino acid analysis of the native peptide and its trypsin and carboxypeptidase fragments indicated that an alanyl residue occupies position 26. This finding is in contrast to the sequence predicted for the beta-lipotropin/corticotropin precursor by recombinant DNA techniques which suggests that the 26th residue of the beta-endorphin molecule should be valine.
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PMID:alpha-N-acetyl-beta-endorphin1-26 from the neurointermediary lobe of the rat pituitary: isolation, purification, and characterization by high-performance liquid chromatography. 684 89

Using the spinal superfusion procedure, in anesthetized rats and cats, the presence of active factors which displace dihydromorphine in brain opiate binding studies, has been observed. Separation of this activity on a Sephadex G-10 column reveals the presence of two fractions which occur before (Fraction I) and after (Fraction II) the salt peak which account for over 70% of the observed dihydromorphine-displacing activity. The ratio of activity in Fraction II/Fraction I is 33 and 21, in the resting spinal perfusates of the rat and cat, respectively. High intensity, bilateral stimulation of the sciatic nerve in cats, results in a 30- and 5.4-fold increase in the levels of Fraction I and Fraction II, respectively, over pre-stimulation levels. In rat, bilateral stimulation of the hind paws, resulted in a frequency-dependent increase in the levels of Fraction I (1.9- and 3.2-fold at 5 and 50 Hz, respectively). Dynorphin 1-13 fragment elutes at least partly in Fraction I. With regard to Fraction II, the peak co-chromatographs with hexapeptide derivatives of enkephalin. Met- and Leu-enkephalin (Fraction III), elute off the column at a point where opiate receptor displacing activity is relatively small. Electrophoretic separation of Fraction I radioreceptor activity of alkaline and acid pH on agarose columns revealed two principle peaks which co-migrated with alpha-neoendorphin and dynorphin 1-13. Fraction II activity appeared primarily in a single peak which was isographic with enkephalin hexapeptides. Using radioimmunoassays, detectable levels of dynorphin and Met-enkephalin were observed and sciatic nerve stimulation resulted in significant increases. Neither column-coupled radioreceptor assays nor radioimmunoassays revealed the presence of beta-endorphin. The present experiments demonstrate the releasability by high intensity somatic stimulation of a variety of opioid peptides present in spinal terminals. Significantly, however, the majority of this activity appears to be found in fractions different from those of the pentapeptide enkephalins.
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PMID:Studies on the release by somatic stimulation from rat and cat spinal cord of active materials which displace dihydromorphine in an opiate-binding assay. 686 Sep 55

N alpha-Acyl amino acid releasing enzyme (NAARE), an enzyme cleaving acetylMet-Ala at the Met-Ala bond was purified from rat brain cytosol to apparent homogeneity by salt precipitation, gel filtration, and several steps of ion exchange. Levels of NAARE exceeded acylase measured with acetylmethionine in all brain regions and subcellular fractions examined: 60% was associated with cytosol and the remainder with debris or the crude nuclear and mitochondrial-synaptosomal subfractions. Activity was highest in pituitary and was approximately 0.5-0.6 that of liver or kidney. The purified enzyme preferentially hydrolyzed acetylmethionyl peptides: Km for acetylMet-Ala was 0.93; Vmax, 3.5 nmol-1 (kcat, 1185) with pH optimum of 8.9 as compared with 8.2 for acylases measured in cytosol. The purified enzyme was devoid of acylase and common exo- and endopeptidase contamination. Structure-activity relationships examined with synthetic formylated or acetylated peptides indicated no significant effects for di- or tripeptides if the second substituent was Ala, Ser, Asn, or Thr, but the activity was reduced 0.5-fold for Leu, a branched-chain amino acid. No hydrolysis was observed for polypeptides with five or more residues having N-terminal acetylated Tyr (enkephalin) or Ser (alpha-melanocyte-stimulating hormone, thymosin alpha 1), supporting the notion that the enzyme plays a role only in turnover of smaller peptides formed perhaps as a result of endopeptidase cleavage of proteins or polypeptides containing acetylated Met at the N terminus.
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PMID:Observations on N alpha-deacetylation of model amino acids and peptides: distribution and purification of a specific N-acyl amino acid releasing enzyme in rat brain. 686 20

Various gastro-entero-pancreatic (GEP) endocrine cells have been shown to contain concomitantly immunoreactivities against several peptide hormones. In the present study the "immunoreactivities" of gastrin (G-) cells of the rat stomach against 21 specific antisera and 10 control sera were investigated by means of the unlabelled antibody enzyme (PAP) technique using modifications of single steps in the immunocytochemical staining sequence. The results indicate that immunoglobulins can bind to gastrin cell granules obviously by non-specific ionic interactions. This non-specific binding of immunoglobulins occurs even in dilution ranges of the sera commonly used in immunohistochemical investigations of the GEP endocrine system. Since "adsorption controls" (preadsorption of the antisera with their respective antigens) will not discriminate between specific and non-specific binding of immunoglobulins to GEP endocrine cells additional specificity controls are necessary. In contrast to the immunostaining of various GEP endocrine cells by "established" antisera and of G-cells by gastrin antiserum immunoglobulins of sera from non-immunized animals as well as antibodies against corticotropin-lipotropin related peptides could be displaced from their binding sites in G-cells by alterations of the NaCl content of the buffers used as diluents or as rinsing solutions. To exclude immunostaining of GEP endocrine cells by nonspecific binding of immunoglobulins the following working procedures are recommended for immunocytochemical investigations of these cells: 1. Use of high titer antisera at low concentrations (diluted 1:1,500 or more). 2. Elevation of the salt (NaCl) content up to 0.5 M of the buffer used as diluent or as rinsing solution. 3. Adsorption controls will show reliable results only if point 1. and 2. have been taken into account.
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PMID:Immunoreactivities of gastrin (G-) cells. II. Non-specific binding of immunoglobulins to G-cells by ionic interactions. 739 Aug 78

A new melanocyte-stimulating peptide has been isolated from acid extracts of frozen human pituitary glands by salt/ethanol fractionation, Sephadex G-75 gel filtration and DEAE- and cM-cellulose ion-exchange chromatography. The peptide is glycosylated, has an N-terminal tryptophan residue and an apparent mol.wt. of 16000 as estimated by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Its amino acid analysis closely resembles residues Trp-105 to Gln-29 predicted for the common precursor protein of bovine corticotropin and beta-lipotropin by Nakanishi, Inoue, Kita, Nakamura, Chang, Cohen & Numa [(1979) Nature (London) 278, 423-427]. This fragment is expected to have melanotropin activity due to the tetrapeptide -His-Phe-Arg-Trp- (residues -51 to -48) of the predicted sequence of the common precursor. It was found to have a molar potency of 1 X 10(-5) relative to alpha-melanotropin in the frog skin bioassay. These characteristics are consistent with the isolated melanotropin peptide being a non-corticotropin, non-lipotropin peptide of the human common precursor protein of corticotropin and lipotropin. The peptide neither potentiates the adrenal weight-maintenance activity of corticotropin-(1-24)-tetracosapeptide when administered to hypophysectomized rats, nor stimulates release of non-esterified fatty acids from isolated rat epididymal cells. A second N-terminal-tryptophan glycopeptide was also isolated, which had an amino-acid composition similar to that predicted for the bovine common precursor protein, residues Trp-105 to Gly-35.
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PMID:Purification and characterization of a gamma-melanotropin precursor from frozen human pituitary glands. 747 89

Proopiomelanocortin (POMC) is a protein that contains the amino acid sequences of numerous peptide hormones, including the melanocyte-stimulating hormones (MSH). MSH peptides of alpha, beta, and gamma primary structure are present in plasma, and all exhibit natriuretic activity. Intravenous infusion of alpha or beta-MSH leads to a time- and dose-dependent natriuresis, whereas gamma-MSH is reported to be natriuretic at low doses but antinatriuretic at high doses. The natriuretic activity of MSH peptides occurs without change in arterial pressure or renal hemodynamics, suggesting a possible direct tubular inhibition of sodium reabsorption. Intravenously infused gamma-MSH is associated with an increase in the plasma concentration of atrial natriuretic peptide. In addition, gamma-MSH also has a direct intrarenal natriuretic action that is dependent on the renal nerves. In rats, gamma-MSH-related peptides are involved in the reflex control of sodium excretion in situations such as the natriuresis that occurs (a) from the remaining kidney after acute unilateral nephrectomy, (b) from the contralateral kidney shortly after unilateral ureteral pressure elevation, and (c) after unilateral carotid artery traction. POMC-derived peptides (including MSH) are modulated in response to salt loading, and alterations in POMC metabolism and plasma peptide concentrations have been observed in genetically hypertensive rats and during the development of adrenal regeneration hypertension. In addition, plasma gamma-MSH levels are elevated in patients with severe congestive heart failure, and in primary hyperaldosteronism. These observations suggest a possible involvement of MSH-related peptides in sodium homeostasis as well as in certain forms of hypertension.
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PMID:Natriuretic properties of melanocyte-stimulating hormones. 750 15


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