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)

Structure-function relationship studies were conducted on the proocytocin/neurophysin endoprotease previously characterized in both bovine neurohypophyseal and corpus luteum granules, using as a reference substrate a synthetic peptide reproducing the entire (1-20) NH2-terminal domain of the precursor. The [D-Arg12] derivative of proocytocin/neurophysin (1-20) was found to be a good competitive inhibitor of the enzyme (Ki = 30 microM), while the [D-Lys11] derivative was not. This allowed the complete purification of two isoforms of the endoprotease (Mr 58,000 and 52,000, respectively) by affinity chromatography using covalently immobilized [D-Arg12] proocytocin/neurophysin (1-20) as the affinity adsorbent. The use of selectively modified or truncated forms of the reference substrate or of the [D-Arg12] competitive inhibitor of the endoprotease established clearly that this basic pair specific convertase is sensitive to modification of the substrate structure either at the basic residues of the cleavage locus or at amino acids around this site (i.e., Pro7 and Gly9). It is concluded that longer distance interactions between amino acids situated on both the NH2 and COOH sides of the basic doublet Lys11Arg12 may contribute to the stabilization of a preferred substrate conformation allowing recognition by the enzyme subsites.
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PMID:Proocytocin/neurophysin convertase from bovine neurohypophysis and corpus luteum secretory granules: complete purification, structure-function relationships, and competitive inhibitor. 265 78

Pro-ocytocin/neurophysin convertase is a divalent cation-dependent endoprotease isolated from both bovine corpus luteum and neurohypophyseal secretory granules. The putative pro-ocytocin/neurophysin converting enzyme cleaves the Arg12-Ala13 bonds of both pro-ocytocin/neurophysin (1----20) and pro-ocytocin/neurophysin obtained by hemisynthesis. The minimal efficient substrate structure allowing recognition by this processing endoprotease was defined by measuring its cleavage efficiency and the inhibitory properties of a set of 34 selectively modified derivatives of the (1----20) NH2-terminal domain of the ocytocin/neurophysin precursor. The data demonstrate that: (i) the basic Lys11-Arg12 doublet, although necessary, is not sufficient; (ii) a minimal substrate length of nine amino acids (residues 7-15 or 8-16) is essential; (iii) those amino acids around the Lys-Arg doublet which contribute to the formation of a possible beta-turn-alpha-helix secondary structure are critical; (iv) substrate recognition by the enzyme may involve several subsites in which structural determinants, situated on both sides of the basic doublet, participate; (v) the NH2-terminal sequence of neurophysin plays a critical role in the correct reading of the cleavage sequence by the processing endoprotease. It is proposed, first, that this type of structural feature may constitute the basis of a general coding system for endoproteases involved in the processing of polypeptide hormone precursors; second, that in addition to its role in the intragranular packaging of the nonapeptide hormone, neurophysin plays a key role in the correct processing of its common precursor with ocytocin.
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PMID:Processing endoprotease recognizes a structural feature at the cleavage site of peptide prohormones. The pro-ocytocin/neurophysin model. 267 20

An enriched preparation of neurosecretory granules from bovine pituitary neural lobes was used as a source of processing enzymes possibly involved in the cleavage of the proocytocin/neurophysin precursor. A synthetic eicosapeptide reproducing the entire (1-20) sequence of the NH2-terminal domain of the bovine ocytocin/neurophysin precursor was used as a substrate to monitor an endoprotease activity cleaving at the Lys11-Arg12 doublet. The 58-kDa endoprotease detected in the lysate of neurohypophyseal granules produced a single cleavage, after the doublet, at the Arg12-Ala13 peptide bond. This endoprotease with pHi 6.9 and 7.2 exhibits maximal activity at pH around neutrality (7.0) and was strongly inhibited by divalent cation chelating agents [ethylenediaminetetraacetic acid and ethylene glycol bis(beta-aminoethyl ether)-N,N,N',-N'-tetraacetic acid] and to some extent by p-(chloromercuri)benzoate and p-(chloromercuri)benzenesulfonic acid, while phenylmethanesulfonyl fluoride and pepstatin were not active. This endoprotease action was sensitive to any modification of the substrate at either basic amino acid of the doublet since replacement of either L-Lys11 or L-Arg12 by D-Lys or D-Arg and by L-Nle abolished the cleavage reaction. In contrast, reversal of the polarity of the doublet in [Arg11,Lys12]proocytocin/neurophysin(1-20) had no effect on the mode of endoproteolytic cleavage as well as modifications of Gly10 (replaced by Ala10). It is concluded that the selectivity of this endoprotease, which may be involved in the primary event occurring in proocytocin/neurophysin processing, is strictly dependent upon the integrity of the basic doublet but that other parameters determined by the amino acid sequence around this doublet may play an important role.
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PMID:Partial purification and functional properties of an endoprotease from bovine neurosecretory granules cleaving proocytocin/neurophysin peptides at the basic amino acid doublet. 282 69

Three putative processing enzymes, each with defined action in a prohormone system, a 'pro-ocytocin-neurophysin convertase' from bovine neurohypophysis secretory granules, a 'Leu-enkephalin Arg6 generating enzyme' from human CSF and the endoprotease from the 'S-28 convertase' complex of rat brain cortex, were tested for their ability to hydrolyze peptides deriving from pro-ocytocin, pro-enkephalin B and pro-somatostatin, respectively at pairs of basic amino acids. The observations suggest that structural parameters specified by the peptide region around the dibasic moieties govern recognition by the enzyme and define which peptide bond is hydrolyzed.
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PMID:Role of peptide substrate structure in the selective processing of peptide prohormones at basic amino acid pairs by endoproteases. 289 32

Two neuropeptide precursor processing enzyme systems were characterized in the rat brain cortex and bovine neurohypophysis and corpus luteum. The first one combines the action of a 90 kDa endoprotease which cleaves somatostatin-28 before the Arg-Lys doublet and that of an aminopeptidase B-like enzyme. The second system associates the action of a 58 kDa endoprotease cleaving pro-ocytocin/neurophysin (1-20) after the Lys-Arg dibasic moiety and a carboxypeptidase B-like activity. Both systems appear to be located in membrane-limited secretory vesicles of the producing organs, and to exhibit the properties of metallo-enzymes sensitive to divalent cation chelators. In contrast, they do not show the characteristics of serine-proteases and of trypsin-like enzymes. Studies with substrate analogs selectively modified at the basic doublet indicated that the integrity of both basic amino acids is essential but that conformational parameters, probably governed by the amino acid sequences flanking the basic doublet, play an important role. These data will be discussed in relation to a hypothesis on the predicted preferred secondary structure of these restriction loci.
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PMID:Somatostatin-28 and pro-ocytocin/neurophysin convertases: basic pair selective endoproteases involved in pro-hormone processing in the rat brain cortex and bovine corpus luteum. 290 27

The selectivity and mechanism of processing at paired basic amino acids in hormone precursors was studied on several analogues of the (1-20)-aminoterminal domain of the ocytocin/neurophysin precursor in a cleavage assay by an endoprotease partially purified from bovine pituitary secretory granules. Peptide analogues with amino acid substitutions in, and around, the basic doublet were synthesized and used as substrates. The data obtained demonstrate the strict requirement of the processing enzyme for basic amino acids in tandem within a possibly preferred conformation which may be highly conserved in the aminoterminal domain of this hormone precursor.
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PMID:Synthetic peptide substrates as models to study a pro-ocytocin/neurophysin converting enzyme. 304 97

The octacosapeptide sequence [Tyr18] pro-ocytocin/neurophysin (1-18)NH2 [pro-OT/Np(1-18)NH2] was synthesized and used as substrate to detect endoprotease(s) possibly involved in the processing of this precursor in bovine hypothalamo-neurohypophyseal tract. An endopeptidase (58 Kda) was detected in Lysates made from highly purified neurosecretory granules. This protease which cleaves the peptide bond on the carboxyl side of the Lys-Arg doublet, and no single basic residue, generates both OT-Gly10-Lys11-Arg12+Ala13-Val-Leu-Asp-Leu-Tyr18 (NH2) from the octacosapeptide substrate. In addition, a carboxypeptidase B-like activity converting OT-Gly10-Lys11-Arg12 into OT-Gly10 was detected in the same granule Lysates. It is hypothesized that a combination of these endoprotease and carboxypeptidase B-like activities together with the amidating enzyme of secretory granules might participate in the cleavage and processing of pro-OT/Np in vivo.
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PMID:An endopeptidase associated with bovine neurohypophysis secretory granules cleaves pro-ocytocin/neurophysin peptide at paired basic residues. 351 14

Purified secretory granules from the corpus luteum of super ovulated and fecundated cows, at day 7-8 after the heat period, were used as a source of pro-ocytocin/neurophysin (pro OT/Np) processing enzymes. An endoprotease comparable to the previously described pituitary enzyme both by its catalytic properties and sensitivity to various inhibitors was characterized. This protease cleaves pro OT/Np (1-20) after the basic Lys11Arg12 doublet to release OT Gly10 Lys11 Arg12. Moreover C-terminally extended ocytocin, i.e. OTGlyLys and OTGlyLysArg together with ocytocin were identified in extracts from the corpus luteum. Together these data argue strongly in favor of pro OT/Np processing pathways in which cleavage of the precursor at the Arg12-Ala13 peptide bond is the primary event.
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PMID:C-terminally extended ocytocin and pro-ocytocin: neurophysin peptide converting enzyme in bovine corpus luteum. 355 57

The activity of pro-oxytocin/neurophysin (pro-OT/Np)-processing enzymes was determined in human granulosa cells, follicular fluids and purified secretory granules of corpora lutea. We detected the presence of an endoprotease which releases OT-Gly10-Lys11-Arg12 on cleavage of the synthetic pro-OT/Np(1-20) peptide after the dibasic Lys11-Arg12 doublet. This endoprotease was inhibited by EDTA, but was not affected by phenylmethanesulfonyl fluoride and pepstatin. Enzymatic activity was markedly reduced by replacement of L-Arg by D-Arg in the basic amino acid doublet of the substrate. The molecular weight of this enzyme was estimated to be 60 kDa. These features closely resembled those of the endoprotease identified in the bovine pituitary. The endoprotease is a metalloenzyme, specific for the Lys-Arg doublet. We also detected a carboxypeptidasic activity, inhibited by guanidinoethane-mercaptosuccinic acid. In the light of our previous detection of the processing intermediates, OT-Gly-Lys-Arg, OT-Gly-Lys and OT-Gly, in human ovary, these observations are in favour of a pro-OT/Np-processing pathway in the human ovary comparable with that in the bovine ovary. Moreover, these results confirm that oxytocin post-translational processing occurs in the human ovary and strongly suggest that pro-OT/Np is synthesized locally.
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PMID:Evidence for the presence of pro-oxytocin/neurophysin-converting enzyme in the human ovary. 793 Oct 7

Proteolytic activation of prohormones and proproteins occurs most frequently at the level of basic amino acids arranged in doublets. Previous predictions by Rholam et al. [Rholam, M., Nicolas, P., & Cohen, P. (1986) FEBS Lett. 207. 1-6] have indicated, on the basis of 20 prohormone sequences containing 53 dibasic potential processing sites, that dibasic sites situated in, or next to, beta-turns were cleaved in vivo, whereas sites included in ordered structures like beta-sheets or alpha-helices were not. We have used peptide analogs of the proocytocin/neurophysin processing domain and a purified preparation of the putative proocytocin convertase from bovine tissues as a model to demonstrate that (1) processing at dibasic sites is associated with a prohormone sequence organized in a beta-turn structure; (2) the beta-turn is an interchangeable motif since the original sequence could be replaced by an heterologous one possessing the ability to organize as a beta-turn; and (3) this particular secondary structure participates in the catalytic reaction, most likely by favoring the interactions of the substrate with the processing endoprotease. It is concluded that, in addition to the dibasic and other amino acids around the cleavage loci, the beta-turn constitutes a key feature in the proteolytic processing reaction in participating as the favorable conformation for optimal substrate-enzyme active site recognition.
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PMID:Role of beta-turn in proteolytic processing of peptide hormone precursors at dibasic sites. 849 28


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