Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01178 (oxytocin)
 15,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Particulate fractions of human small intestinal mucosa contain an enzyme capable of hydrolyzing N-benzoyl-L-tyrosyl-p-aminobenzoic acid (PABA-peptide), a substrate used for clinical purposes to assess exocrine function of the pancreas (PABA test, pancreas function test). In this paper we describe the purification of PABA-peptide hydrolase (PPH) by immunoaffinity chromatography using a monoclonal antibody (Mab), HBB 3/716/36, bound to protein A-Sepharose, and the characterization of the purified enzyme. The final preparation of the enzyme was in the immobilized form, i.e., bound to Mab-protein A-Sepharose, and showed a 765-fold enrichment over the mucosal homogenate. The enrichment factor in purified microvillus membranes was comparable to that of sucrase-isomaltase, a microvillar marker enzyme. This, together with immunoelectron microscopy using protein A-gold, indicated that PPH is located in the apical membrane of intestinal epithelial cells. The enzyme was found to be present throughout the small intestine with the activity in distal ileum being 4.5-fold higher than that in the proximal duodenum. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the immunoaffinity-purified PPH under reducing conditions revealed a polypeptide band with a relative molecular weight (Mr) of 100,000; under nonreducing conditions a major band with Mr 200,000 was observed. This indicates that PPH consists of two subunits with Mr 100,000 each, which are held together by one or more disulfide bonds. Two-dimensional polyacrylamide gel electrophoresis of the enzyme showed marked microheterogeneity, with pI's ranging from 6.0 to 6.85, probably due to glycosylation. The Km for PABA-peptide was 16.7 mM, and the pH optimum was 7.5-8.0 PPH activity was not inhibited by phenylmethylsulfonyl fluoride; pepstatin, leupeptin, amastatin, bestatin, puromycin, iodoacetate, or phosphoramidon. Activity was affected by captopril and Zinkov inhibitor, and in particular by thiol and chelating reagents. Chelator-inhibited PPH could be reactivated by bivalent metal ions, Zn2+ being the most effective. The enzyme catalyzed the hydrolysis of peptides including insulin B-chain, angiotensins I and II, bradykinin and bradykinin derivatives, oxytocin, and substance P, in each case yielding reproducible peptide fragments. On the basis of amino acid analysis of the products it could be concluded that peptides are hydrolyzed preferentially after an aromatic residue.
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PMID:N-benzoyl-L-tyrosyl-p-aminobenzoic acid hydrolase: a metalloendopeptidase of the human intestinal microvillus membrane which degrades biologically active peptides. 326 61

Post-proline endopeptidase (PPE, EC 3.4.21.26) was purified 3,450 times from human lung. PPE was routinely assayed with the artificial substrate, carbobenzoxy-glycyl-L-prolyl-p-nitroanilide (Z-Gly-Pro-pNA). The pH optimum was 7.4, and the Mr was 77,000. Thiol blocking agents were strongly inhibitory but serine blocking agents were not inhibitory. No metal ions were required for activity, but heavy metal ions such as Hg2+, Cu2+, Cd2+, and Zn2+ completely inactivated the enzyme. Both dithiothreitol (DTT) and ethylenediaminetetraacetic acid (EDTA) were required to stabilize PPE activity. Michaelis constant values for Z-Gly-Pro-pNA and carbobenzoxy-glycyl-L-prolyl-2-naphthylamide were 0.36 and 0.10 mmol/l, respectively. PPE cleaved vasoactive peptides including bradykinin (BK) and des-(Arg9)-BK (Pro3-Gly4 and Pro7-Phe8 bonds), angiotensins I and II (Pro7-Phe8 bond), substance P (Pro4-Gln5 bond), and oxytocin (Pro7-Leu8 bond). Each of these peptides inhibited PPE-catalyzed hydrolysis of Z-Gly-Pro-pNA competitively. BK had the lowest Ki value (2.35 mumol/l) and oxytocin had the highest Ki value (84.0 mumol/l). PPE was not inhibited by captopril, a potent inhibitor of angiotensin converting enzyme, which also cleaves the Pro7-Phe8 bond of BK.
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PMID:Human lung post-proline endopeptidase: purification and action on vasoactive peptides. 354 26

Nulliparous female Sprague-Dawley rats, cannulated in the left lateral ventricle, were ovariectomized and estrogen primed, then either rendered anosmic via intranasal irrigation with zinc sulfate or left with intact olfaction. Forty-eight hr later, after a 2-hr habituation to the test cage, these animals were injected with either intracerebroventricular oxytocin (400 ng in 2 microliter saline) or saline (2 microliter). Only the group receiving both zinc sulfate and oxytocin became maternal. Additionally, approximately one third of the olfaction-intact rats and none of the anosmic rats cannibalized the rat pups. These results are discussed in regard to discrepancies in the literature regarding oxytocin's role in inducing maternal behavior, as well as the functional connection of the olfactory and oxytocin systems.
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PMID:The ability of oxytocin to induce short latency maternal behavior is dependent on peripheral anosmia. 360 16

Changes in the mineral composition of mare milk during lactation were studied. Milk samples were obtained from five Thoroughbred mares one to three times weekly from the first to the eighth week of lactation and from two of the mares for an additional 8 wk. Samples averaging 500 mL were obtained after oxytocin was administered to the mares. Each sample was analyzed for total solids, ash, calcium, phosphorus, magnesium, sodium, potassium, copper and zinc. The concentration of all constituents except sodium and potassium decreased throughout lactation. The rates of decline of ash, calcium, phosphorus and magnesium concentration were similar, but the rates of decline of the other elements differed. Thus, the mineral composition of mare milk should be described in terms of the stage of lactation of the mare. The total solids and ash content of mare milk were 12 and 0.61% respectively, at the end of the first week of lactation, 10.5 and 0.45% at 4 wk, 10 and 0.38% at 8 wk and 10.2 and 0.32% at 16 wk. The calcium, phosphorus and magnesium concentrations at the end of the same periods were 1345, 943 and 118 micrograms/g of milk at 1 wk; 1070, 659 and 86 at 4 wk; 831, 574 and 58 at 8 wk and 700, 540 and 43 micrograms/g of milk at 16 wk. Copper and zinc concentrations were 0.85 and 3.1, 0.55 and 2.2, 0.29 and 1.9 and 0.28 and 1.8 microgram/g of milk at 1, 4, 8 and 16 wk, respectively.
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PMID:Lactation in the horse: the mineral composition of mare milk. 379 22

Previously it has been shown that vasopressin (VP) and oxytocin are converted by aminopeptidase activity in brain membranes into fragments with potent CNS activities. This report concerns the properties of this enzyme activity, addressed as VP-converting aminopeptidase (VP-AP) activity, in membranes of the rat brain. The VP-AP activity had a pH optimum at pH 7.0 and had a Km of 17 microM for its action on VP. Amastatin was the most potent aminopeptidase inhibitor. Enzyme activity was inhibited by relatively low concentrations of metal chelators. Treatment of brain membranes by EDTA resulted in loss of enzyme activity that was completely reversed by 10 microM Zn2+, indicating that VP-AP activity is a metallopeptidase. Several VP analogues and fragments, in particular VP(1-8), inhibited the action of enzyme activity on VP. Among peptides unrelated to VP, angiotension I, somatostatin, and porcine ACTH(1-39) markedly inhibited enzyme activity. Solubilization of VP-AP activity from brain membranes and gel filtration on Sephadex G200 showed two peaks of activity, one eluting with an apparent mass of about 140 kDa, the other in the void volume. Gel filtration fractions were able to convert [3H][Phe3]VP in a step-wise fashion. The VP-AP-like activity was found in many tissues outside the brain. Highest activity was present in lung, kidney, parts of the gastrointestinal tract, ovary, and uterus. The results indicate that VP-AP activity is a widely distributed enzyme with probably multiple functions, one of which involves the metabolism of vasopressin in the brain.
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PMID:Properties of aminopeptidase activity involved in the conversion of vasopressin by rat brain membranes. 799 91

The serum level of placental leucine aminopeptidase (P-LAP) increases during pregnancy. P-LAP degrades several peptide hormones such as oxytocin and vasopresin, suggesting a role in maintaining homeostasis during pregnancy. In the study reported here, we have isolated a cDNA clone with 4084 base pairs encoding P-LAP from a human placental cDNA library. The amino acid sequence deduced from the cDNA contained all of the sequences of the peptide fragments obtained by digestion of the purified protein with trypsin. The predicted P-LAP contains the HEXXH consensus sequence of zinc metallopeptidases, indicating that the enzyme belongs to this family, which includes aminopeptidase N and aminopeptidase A. The deduced sequence also contains a hydrophobic region near the N terminus, suggesting that the enzyme is a type II integral membrane protein. Northern blot analysis revealed that P-LAP was expressed in several tissues, some of which expressed two forms of mRNAs. These results suggest that the enzyme is synthesized as an integral membrane protein and is released into blood under some physiological conditions.
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PMID:Human placental leucine aminopeptidase/oxytocinase. A new member of type II membrane-spanning zinc metallopeptidase family. 855 Jun 19

Recent evidence suggests that the gas nitric oxide can modulate the secretion of a number of hypothalamic hormones, and may be co-localized particularly to oxytocin-containing neurons. Another gas, carbon monoxide (CO), has also been suggested to play a role in neural signaling in the brain, and the synthetic enzyme responsible for the generation of carbon monoxide has been reported to be present in the rat hypothalamus. In this study, we have therefore investigated whether CO has the ability to modify the release of oxytocin from acute rat hypothalamic explants. Hemin, a specific CO precursor through the enzyme heme oxygenase (the enzymatic pathway synthesizing endogenous CO, was found to inhibit KCl-stimulated oxytocin release, with a maximal effect at 10(-5) M, while showing no effect on basal oxytocin secretion. The stimulation of oxytocin by serotonin 10 ng/ml was also significantly antagonized by hemin 10(-7) M. An inhibitor of heme oxygenase, zinc-protoporphyrin-9, had no effect on basal or stimulated oxytocin release. When hemin and zinc-protoporphyrin-9 were given together, the hemin-induced inhibition of oxytocin was completely antagonized by the enzyme inhibitor. Ferrous hemoglobin A0, a substance known to bind CO with high affinity, had no effect on either basal or stimulated oxytocin release, but when hemin and ferrous hemoglobin A0 were given together the hemin-induced inhibition of oxytocin was completely blocked. These findings provide evidence that endogenous CO may play a role in the control of oxytocin release and that, by analogy with nitric oxide, CO may represent a major new neuroendocrine modulator.
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PMID:Oxytocin release is inhibited by the generation of carbon monoxide from the rat hypothalamus--further evidence for carbon monoxide as a neuromodulator. 901 87

Serum levels of human placental leucine aminopeptidase/oxytocinase (P-LAP) increase with gestation. cDNA cloning of P-LAP revealed that the enzyme is a type II membrane-bound protein containing the consensus HEXXH(X)18E motif found in the M1 family of zinc-metallopeptidase proteins. In this study, a recombinant soluble form of P-LAP found in maternal serum was expressed in Chinese hamster ovary cells, purified to homogeneity and then characterized. Although N-terminal sequencing revealed a four-amino-acid deletion, the purified enzyme was active and was shown to be a zinc-containing homodimeric protein with molecular mass of 280 kDa in solution. Using artificial substrates, it was shown that the enzyme has broad specificity and is inhibited by several compounds known as aminopeptidase inhibitors. Subsequently, sequential N-terminal amino-acid liberation of several peptide hormones by the enzyme was monitored and structures of the products were determined. Among the hormones having a cysteine residue at their N-terminal end and intramolecular disulfide bonds, it was found that vasopressin and oxytocin, but not calcitonin and endothelins, were cleaved by the enzyme. Because the molecular properties of oxytocinase so far reported often conflict, our results provide an initial biochemical and enzymatic characterization of moleculary defined P-LAP/oxytocinase.
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PMID:Characterization of a recombinant soluble form of human placental leucine aminopeptidase/oxytocinase expressed in Chinese hamster ovary cells. 1060 49

The insulin-regulated membrane aminopeptidase (IRAP) was originally identified in fat and muscle cells as a major protein in intracellular vesicles that also harbor the insulin-responsive glucose transporter GLUT4. IRAP, like GLUT4, predominantly localizes to these intracellular vesicles under basal conditions. In response to insulin IRAP, like GLUT4, translocates to the plasma membrane. Purification and cloning of IRAP revealed that it was a novel member of the family of zinc-dependent membrane aminopeptidases. Upon the cloning of the human placental oxytocinase (P-LAP) it was discovered that IRAP and P-LAP were the rat and human homologues of the same protein. The expression of IRAP/P-LAP is not limited to fat and muscle cells, and the subcellular distribution of IRAP/P-LAP is regulated by different peptide hormones and exercise. IRAP/P-LAP cleaves several peptide hormones in vitro. In insulin- and oxytocin-treated cells, concomitant with the appearance of IRAP/P-LAP at the cell surface, aminopeptidase activity toward extracellular substrates increases. A physiological function for IRAP/P-LAP may thus be the processing of circulating peptide hormones. These extracellular substrates, however, would be processed efficiently only when IRAP/P-LAP gets access to them after translocation to the cell surface upon stimulation of cells with insulin or other factors. The in vivo substrates for IRAP/P-LAP remain to be determined. The initial characterization of mice in which IRAP/P-LAP was deleted (IRAP -/- mice) revealed that GLUT4 protein levels were dramatically decreased in all fat and muscle tissues. This finding suggests a function for IRAP/P-LAP in the regulation of GLUT4 levels. Further characterization of the IRAP -/- mice is required to elucidate the role IRAP/P-LAP may play in the control of peptide hormone metabolism.
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PMID:The insulin-regulated aminopeptidase: a companion and regulator of GLUT4. 1270 Jan

The emergence of new technologies from the genomics revolution will transform the potential application of biomarkers to assess how pollutants impact people, animals, and ecosystems. Genetic databases provide a huge resource from which candidate molecular biomarkers can be identified and, subsequently, exploited to address these issues. However, a major challenge is to link these novel molecular indices to ecologically relevant whole-organism life-cycle traits (such as reproduction and growth). Such a functional link is provided by annetocin, previously characterized as a member of the vasopressin/oxytocin superfamily of neuropeptides. It is expressed in annelid worms within the neurons of the central nervous system and has been shown to be involved in the induction of egg-laying behavior. This paper outlines the validation of annetocin as a novel biomarker of reproductive fitness in the earthworm Eisenia fetida. The design of primer pairs targeted toward oligochaete annetocin has facilitated the isolation of a full-length annetocin cDNA from this species. Optimization of a real-time quantitative PCR procedure exploiting the fluorescent DNA-binding molecule, Sybr Green, has allowed the measurement of annetocin transcript levels over a range covering six orders of magnitude. Using this approach, gene expression was measured in earthworms exposed to soils polluted with high concentrations of zinc and lead. Traditional growth and reproductive indices, including cocoon production, were also recorded and related to the molecular parameter. The future use of annetocin as a molecular genetic biomarker in terrestrial ecotoxicology is discussed.
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PMID:Measurement of annetocin gene expression: a new reproductive biomarker in earthworm ecotoxicology. 1465 61


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