Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P20366 (substance P)
 21,176 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have recently shown that (a) [125I-Tyr8]bradykinin (BK) recognized bradykinin binding sites in guinea pig epithelium membranes with a Kd value of 1.6 nM and a Bmax of 156 fmol/mg protein, and (b) B2 agonists and some B2 antagonists, such as D-Arg-[Hyp3,D-Phe7,Leu8]BK, inhibited this specific binding with a Ki value of 32 nM. In the present study, we have radioiodinated the B2 antagonist Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]BK and have performed a full characterization of the binding properties of this tracer in the same membrane preparation. Equilibrium experiments performed in the absence or presence of an excess of BK (10(-5) M) showed that 125I-Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]BK specifically labelled two different sites. One of these is the same as the site labelled by [125I-Tyr8]BK, and this indicates that 125I-Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]BK interacts specifically with kinin B2 receptors. Equilibrium experiment performed in the presence of an excess of BK (10(-5) M) indicated that specific binding of 125I-Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]BK to the second site is also saturable and Scatchard analysis showed that the site is of high affinity with a Kd of 16.8 nM and a Bmax of 2.08 pmol/mg protein. Surprisingly, unlabelled B2 agonists such as bradykinin, [Tyr8]BK, [Leu8]BK, [Hyp3,Tyr8(OMe)]BK, D-Arg-[Hyp3]BK and kallidin were found to be inactive on this second site. A series of B2 receptor antagonists, Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]BK, D-Arg-[Hyp3,D-Phe7,Leu8]BK, D-Arg-[Hyp3,Leu5,8,D-Phe7]BK, D-Arg-[Hyp3,Gly6,D-Phe7,Leu8]BK and D-Arg-[Hyp3,Thi5,8,D-Phe7]BK inhibited 125I-Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]BK binding with Ki values of 25.0, 20.9, 15.8, 64.6 and 6606.9 nM respectively. On the other hand, [Thi5,8,D-Phe7]BK did not interfere with 125I-Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]BK but was found to be a potent inhibitor of [125I-Tyr8]BK binding (Ki = 53.7 nM). As expected, B1 receptor agonists, antagonists and peptides non-related to BK such as substance P, neurokinin A, neurokinin B, angiotensin II, bombesin, vasopressin and the calcitonin gene related peptide were unable to compete with 125I-Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]BK. The results show that 125I-Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]BK is interacting with two distinct binding sites in the guinea pig epithelium: one is the well known bradykinin B2 receptor and the other is a new, non-characterized binding site that interacts exclusively with some bradykinin receptor antagonists.
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PMID:Characterization of a novel binding site for 125I-Tyr-D-Arg-[Hyp3,D-Phe7,Leu8]bradykinin on epithelial membranes of guinea pig ileum. 132 66

The effect of opioids on the release of immunoreactive substance P (iSP) following simultaneous electrical stimulation of the sectioned sciatic and saphenous nerves was examined by perfusion of the subcutaneous space in the rat instep. Antidromic stimulation of both the nerves caused an increase in iSP release, which was dependent on the intensity of stimulation, and an approx. 200% increase in Evans blue extravasation. Stimulation-induced iSP release and extravasation were suppressed by pretreatment with capsaicin (50 mg/kg s.c.) and spantide (10 mumol/kg i.p.), respectively. Intra-arterial infusion of morphine (30 mumol/kg) or ethylketocyclazocine (30 mumol/kg) or [D-Ala2,D-Leu5]enkephalin (30 mumol/kg) inhibited the increase in iSP release evoked by antidromic stimulation at 10 V. This inhibitory effect of morphine was antagonized by pretreatment with naloxone (2 mg/kg, i.p.). These results suggest existence of multiple types of opioid receptor on the peripheral endings of primary afferent fibers, that regulate SP release from the peripheral nerve endings into the extravascular space.
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PMID:Influence of opioids on substance P release evoked by antidromic stimulation of primary afferent fibers in the hind instep of rats. 137 91

Neutral endopeptidase 24.11 contains an active site arginine believed to function in substrate binding. This arginine is thought to form an ionic interaction with the COOH-terminal carboxylate of NEP substrates. The functionality of arginine 102 has been investigated by using site-directed mutagenesis to produce mutants in which this residue was converted to a lysine, glycine, glutamine, or glutamate. All of the mutants exhibited essentially full activity as determined with a synthetic peptide amide, glutaryl-Ala-Ala-Phe-4-methoxy-2-naphthylamide. In contrast, activity was detected only with the wild-type enzyme and the lysine mutant using a synthetic substrate containing a free COOH-terminal carboxylate, dansyl-Gly-Trp-Gly. Inhibition studies with the physiologically active peptide substrates substance P, endothelin, and angiotensin I, as well as substance P free acid, [D-Ala2,Leu5]enkephalin, and [D-Ala2,Leu5]enkephalinamide indicated a lack of importance of arginine 102 in substrate binding. With [D-Ala2,Met5]enkephalin and the chemotactic peptide, N-formyl-Met-Leu-Phe, a significant decrease in affinity is observed with the arginine 102 mutants. These results suggest that the contribution of arginine 102 to substrate binding is dependent upon the strength of other subsite interactions. Examination of dipeptides as inhibitors indicates that the nature and orientation of the P'2 residue is important in determining the strength of the interaction of arginine 102 with its substrates.
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PMID:Analysis of the importance of arginine 102 in neutral endopeptidase (enkephalinase) catalysis. 137 21

Kinins and substance P have been implicated in the pathogenesis of inflammatory arthritis by virtue of their abilities to induce vasodilation, edema, and pain. The relative biological potencies of these peptides in vivo would depend at least in part upon their rates of catabolism in the joint. We hypothesized that human synovial lining cells may regulate intraarticular levels of kinins and neuropeptides via degradation by cell surface-associated peptidases. We exposed intact human synovial fibroblasts to kinins and substance P, in the presence or absence of specific peptidase inhibitors, and measured the amount of intact substrate remaining and degradation product(s) generated over time. Aminopeptidase M (AmM; EC 3.4.11.2), neutral endopeptidase-24.11 (NEP-24.11; EC 3.4.24.11), and dipeptidyl(amino)peptidase IV (DAP IV; EC 3.4.14.5) were identified on the cell surface of synovial cells. Bradykinin degradation was due entirely to NEP-24.11 (1.39 +/- 0.29 nmol/min per well). Lysylbradykinin was also degraded by NEP-24.11 (0.80 +/- 0.19 nmol/min per well); however, in the presence of phosphoramidon, AmM-mediated conversion to bradykinin (3.74 +/- 0.46 nmol/min per well) could be demonstrated. The combined actions of NEP-24.11 (0.93 +/- 0.15 nmol/min per well) and DAP IV (0.84 +/- 0.18 nmol/min per well) were responsible for the degradation of substance P. AmM (2.44 +/- 0.33 nmol/min per well) and NEP-24.11 (1.30 +/- 0.45 nmol/min per well) were responsible for the degradation of the opioid peptide, [Leu5]enkephalin. The identity of each of the three peptidases was confirmed via synthetic substrate hydrolysis, inhibition profile, and immunological identification. The profiles of peptidase enzymes identified in cells derived from rheumatoid and osteoarthritic joints were identical. These data demonstrate the human synovial fibroblast to be a rich source of three specific peptidases and suggest that it may play a prominent role in regulating peptide levels in the joint.
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PMID:Cultured human synovial fibroblasts rapidly metabolize kinins and neuropeptides. 138 26

The effect of a unilateral 6-hydroxydopamine (6-OHDA) lesion of the medial forebrain bundle or of a sham lesion on the neuropeptide content of the striatum and substantia nigra was investigated with or without 6 months L-3,4-dihydroxyphenylalanine (L-DOPA; 200 mg/kg per day) plus carbidopa (25 mg/kg per day) treatment. [Met5]- and [Leu5]enkephalin, substance P (SP), neurotensin (NT) and cholecystokinin (CCK) were measured by a combined HPLC/RIA method. Neurotensin levels were increased in the striatum, and [Leu5]enkephalin, and SP levels were reduced in the substantia nigra as a consequence of the lesion, while the levels of other peptides were unaltered. Administration of L-DOPA to sham-operated rats bilaterally increased SP levels in striatum and substantia nigra, and [Met5]enkephalin and CCK content in substantia nigra. L-DOPA treatment of 6-OHDA-lesioned rats increased [Met5]- and [Leu5]enkephalin and CCK levels in the striatum ipsilateral to the lesion but not on the intact side. In the substantia nigra, the lesion-induced decrease in [Leu5]enkephalin and SP was reversed by L-DOPA treatment, [Met5]enkephalin and CCK levels ipsilateral to the lesion were further enhanced, and there was an increase in NT ipsilateral to the lesion. Cryptic [Met5]- and [Leu5]enkephalin increased in the ipsilateral striatum following an 6-OHDA lesion. L-DOPA treatment did not alter cryptic enkephalin levels or the lesion-induced increase in cryptic [Met5]enkephalin, while cryptic [Leu5]enkephalin was further increased in lesioned animals given L-DOPA. These results suggest that the pattern of change in basal ganglia peptides in Parkinson's disease is not due solely to the destruction of the nigrostriatal pathway, the drug treatment of the disease or a combination of these factors.
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PMID:Effects of a unilateral 6-hydroxydopamine lesion and prolonged L-3,4-dihydroxyphenylalanine treatment on peptidergic systems in rat basal ganglia. 138 71

We discovered an enzyme in human platelets that deamidates substance P and other tachykinins. Because an amidated carboxyl terminus is important for biological activity, we purified and characterized this deamidase. The enzyme, released from human platelets by thrombin, was purified to homogeneity by ammonium sulfate precipitation, followed by chromatography on an octyl-Sepharose column and chromatofocusing on PBE 94. The purified enzyme exhibits esterase, peptidase, and deamidase activities. The peptidase activity (with furylacryloyl-Phe-Phe) is optimal at pH 5.0 while the esterase (benzoyl-tyrosine ethyl ester) and deamidase (D-Ala2-Leu5-enkephalinamide) activities are optimal at pH 7.0. With biologically important peptides, the enzyme acts both as a deamidase (substance P, neurokinin A, and eledoisin) and a carboxy-peptidase (with bradykinin, angiotensin I, substance P-free acid, oxytocin-free acid) at neutrality, although the carboxypeptidase action is faster at pH 5.5. Enkephalins, released upon deamidation of enkephalinamides, were not cleaved. Gly9-NH2 of oxytocin was released without deamidation. Peptides with a penultimate Arg residue were not hydrolyzed. Some properties of the deamidase are similar to those reported for cathepsin A. The deamidase is inhibited by diisopropylfluorophosphate, inhibitors of chymotrypsin-type enzymes, and mercury compounds while other inhibitors of catheptic enzymes, trypsin-like enzymes, and metalloproteases were ineffective. In gel filtration, the native enzyme has an Mr = 94,000 while in non-reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis the Mr = 52,000 indicating it exists as a dimer. After reduction, deamidase dissociates into two chains of Mr = 33,000 and 21,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. [3H]diisopropylfluorophosphate labeled the active site serine in the Mr = 33,000 chain. The first 25 amino acids of both chains were sequenced. They are identical with the sequences of the two chains of lysosomal "protective protein" which, in turn, has sequence similarity to the KEX1 gene product and carboxypeptidase Y of yeast. This protective protein complexes with beta-galactosidase and neuraminidase in lysosomes and is vitally important in maintaining their activity and stability. A defect in this protein is the cause of galactosialidosis, a severe genetic disorder. The ability of physiological stimuli (e.g. thrombin or collagen) to release the deamidase from platelets indicates that it may also be involved in the local metabolism of bioactive peptides.
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PMID:A peptidase in human platelets that deamidates tachykinins. Probable identity with the lysosomal "protective protein". 169 76

The substituted glucopyranose ring structure 2-hydroxypropyl-beta-cyclodextrin (CDEX) increases the solubility of molecules by inclusion of the agent in the lipophilic interior of the ring. This property is of particular use for the administration of molecules by the intracerebral (ICV) or intrathecal (IT) routes. In concentrations up to 40% w/v (isotonic), this agent (10 microliters) effect upon nociceptive or motor function after IT injection or on EEG and general behavior after ICV injection in rats. Using 20% CDEX, there is no change in the ED50 as compared to saline on the hot plate (HP) after IT injection of morphine, D-Ala2-D-Leu5 enkephalin or Tyr-Aib-Gly-gPhe-mAib-NH2, (Aib: alpha-aminoisobutyric acid) although there is an increase in their respective durations of effect. Cyclic peptide opioids: Tyr-c[D-A2bu-Gly-D-beta Nal(1)-D-Leu] (A2bu: alpha, gamma-diaminobutyric acid; beta-Nal(1): beta-naphthylalanine(1)) or Tyr-c[DA2bu-Gly-beta Nal(1)-D-Leu] are insoluble in saline but are readily dissolved in CDEX, and display a naloxone-sensitive antinociception following spinal administration. In other studies, saline insoluble capsaicin is administered in 25% dimethylsulfoxide (DMSO) or 20% CDEX (15 microliters; 5 mg/ml) which result in a significant reduction in the spinal levels of substance P and calcitonin gene related peptide and an increase in the HP latency. DMSO alone, but not CDEX alone, reduces the levels of the two peptides. These data emphasize the utility of complexation with CDEX for intracerebral drug delivery and compatibility with brain and spinal tissue.
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PMID:The utility of 2-hydroxypropyl-beta-cyclodextrin as a vehicle for the intracerebral and intrathecal administration of drugs. 170 20

The isosteric methyleneoxy psi (CH2O) function was employed as a novel peptide-bond surrogate and incorporated into sequences of two neuropeptides, substance P (SP) and enkephalin. A pseudopeptide analogue [pGlu6,Phe8 psi(CH2O)Gly9]SP6-11 (7) of SP related C-terminal hexapeptide [pGlu6]SP6-11 and two pseudopeptide analogues of [Leu5]enkephalinamide, [Tyr1 psi (CH2O)Gly2, Leu5] enkephalinamide (11) and [Gly2 psi (CH2O)-Gly3, Leu5]enkephalinamide (17), were synthesized. The N alpha-protected pseudodipeptidic units were incorporated in the appropriate peptide sequences by using conventional coupling methods in solution. Compound 7 was a potent agonist (EC50 = 4.8 nM) of substance P as compared to the parent peptide [pGlu6]SP6-11 (EC50 = 1.2 nM), in stimulating contraction of the isolated guinea pig ileum (GPI). Analogue 7 was more potent on the neuronal (NK-3) than on the muscular (NK-1) tachykinin receptors in the GPI as shown by the ratio of activities, EC50 (NK-1)/EC50 (NK-3) = 3.16, thus displaying an improved selectivity for the NK-3 tachykinin receptor subtype as compared to that of [pGlu6]SP6-11, EC50 (NK-1)/EC50 (NK-3) = 0.44. In the rat vas deferens (RVD) assay, a typical NK-2 system, the pseudopeptide analogue 7 was (EC50 = 2 microM) 10-fold more potent than the parent peptide and 20-fold less potent than eledoisin, an NK-2 selective tachykinin. The pseudopeptide enkephalin analogue 17 had low biological activity when tested in the electrically induced GPI (EC50 = 2.3 microM) and was inactive in the mouse vas deferens (MVD) assay. In the rat brain membrane (RBM) binding assay analogue 17 had low affinity (in the micromolar range) for both the mu and delta binding sites. In contrast, analogue 11 was a potent enkephalin agonist (EC50 = 30 nM), being equipotent to [D-Ala2, Leu5]enkephalinamide (DALE) in the GPI assay. In the MVD, analogue 11 showed a substantially reduced activity (EC50 = 92 nM), being about 10-fold less potent than DALE. In the RBM binding assay analogue 11 showed high affinity (in the nanomolar range) for both mu and delta binding sites with increased selectivity for the delta sites as shown by the ratio of the apparent affinities for both receptors, Ki (delta)/Ki (mu) = 2.1. The contribution of the modified peptide bonds in the mode of interaction of SP and enkephalin at their corresponding receptors is discussed.
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PMID:Pseudopeptide analogues of substance P and leucine enkephalinamide containing the psi (CH2O) modification: synthesis and biological activity. 171 57

Aged common marmosets were treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP; 0.5-2.0 mg/kg/week i.p.) for 16 or 24 weeks, observed for a total of 30 weeks and then killed for measurement of biochemical parameters in basal ganglia. The MPTP treatment induced a marked depletion in dopamine, 3,4-dihydroxyphenylacetic acid and homovanillic acid levels in the caudate nucleus and putamen. In contrast, the concentrations of five neuropeptides: [Met5]-enkephalin, [Leu5]-enkephalin, cholecystokinin, substance P and neurotensin as measured by a combined HPLC/RIA method, remained unaltered in all basal ganglia regions examined. Enkephalin precursor levels, as reflected by cryptic [Met5]-enkephalin content, were increased in the putamen, but not in the caudate nucleus, as a consequence of MPTP administration. Cryptic [Leu5]-enkephalin content remained unchanged in the striatum of MPTP treated marmosets. Overall, these results suggest an increase in striatal [Met5]-enkephalin release following chronic MPTP treatment of aged marmosets. However, the chronic treatment of aged marmosets with MPTP does not reproduce the neuropeptide alterations characteristic of Parkinson's disease.
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PMID:Neuropeptide levels in the basal ganglia of aged common marmosets following prolonged treatment with MPTP. 171 7

To investigate the participation of neuropeptides present in the peripheral endings of primary afferent neurons in the inflammatory response, immunoreactive substance P (iSP), calcitonin gene-related peptide (iCGRP) and neurokinin A (iNKA) levels in the s.c. perfusate, and inflammatory response (edema and plasma protein extravasation) evoked in rat paw by noxious stimulation were determined. The effects of these peptides on plasma protein extravasation in the skin of the hind paw of mice were also examined with the pontamine sky blue protein labelling method. The following results were obtained. 1) Immersion of the rat hind paw for 30 min into hot water adjusted to 47 degrees C led to a marked increase in the release of iSP and iCGRP in the subcutaneous perfusate with the formation of thermal edema. 2) Mechanical stimulation (600 g, 10 min) to the hind paw or electrical stimulation of the saphenous and sciatic nerves (10 V, 2 Hz, 1msec duration, 10 min) evoked the increase of iSP release in the perfusate with plasma protein extravasation. 3) iNKA release was not affected by neither heat nor mechanical stimulation. 4) Intraplantar injection of SP, CGRP and NKA induced plasma protein extravasation, the order of potencies being SP greater than CGRP greater than NKA. The action of SP was antagonized by spantide, an SP antagonist. The injection of CGRP with SP produced a synergistic action on plasma protein extravasation. 5) Neonatal pretreatment with capsaicin, which is known to degenerate small-diameter primary afferent neurons, caused the decrease in amount of iSP and iCGRP released during noxious heat stimulation. 6) Pretreatment with Compound 48/80, or stem bromelain and emorphazone, or des-Arg9-[Leu8]-BK, inhibited the iSP release evoked by noxious heat stimulation. 7) Opioids such as morphine (mu-agonist) and ethylketocyclazocine (kappa agonist) inhibited the heat stimulus-evoked iSP release and thermal edema, and the inhibitory effects were antagonized by pretreatment with their antagonists. 8) Morphine or ethylketocyclazocine or [D-Ala2,D-Leu5]-enkephalin (delta-agonist) inhibited the release of iSP evoked by electrical stimulation of the saphenous and sciatic nerves. These results indicate that SP and CGRP present in peripheral endings of small-diameter primary afferent neurons play an important role in the inflammatory response, and that opioids are involved in the regulation of inflammatory response through the inhibition of SP release.
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PMID:[A pharmacological study of the participation of the peripheral endings of primary afferent neurons in the inflammatory response evoked by heat and mechanical noxious stimulation]. 172 88


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