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1.
The opioid peptide H‐Tyr‐c[D‐Cys‐Phe‐Phe‐Cys]NH2 cyclized via a methylene dithiother is a potent and selective μ opioid agonist (Przydial M.J. et al., J Peptide Res, 66, 2005, 255). Dicarba analogues of this peptide with Tyr, 2′,6′‐dimethyltyrosine (Dmt), 3‐[2,6‐dimethyl‐4‐hydroxyphenyl)propanoic acid (Dhp) or (2S)‐2‐methyl‐3‐(2,6‐dimethyl‐4‐hydroxyphenyl)propanoic acid [(2S)‐Mdp] in the 1‐position were prepared. The peptides were synthesized on solid‐phase by substituting d ‐allylglycine and (2S)‐2‐amino‐5‐hexenoic acid in position 2 and 5, respectively, followed by ring‐closing metathesis. Mixtures of cis and trans isomers of the resulting olefinic peptides were obtained, and catalytic hydrogenation yielded the saturated –CH2–CH2– bridged peptides. All six Tyr1‐ and Dmt1‐dicarba analogues retained high μ and δ opioid agonist potency and showed only slight or no preference for μ over δ receptors. As expected, the six Dhp1‐ and (2S)‐Mdp1‐dicarba analogues turned out to be μ opioid antagonists but, surprisingly, displayed a range of different efficacies (agonism, partial agonism or antagonism) at the δ receptor. The obtained results indicate that the μ versus δ receptor selectivity and the efficacy at the δ receptor of these cyclic peptides depend on distinct conformational characteristics of the 15‐membered peptide ring structure, which may affect the spatial positioning of the exocyclic residue and of the Phe3 and Phe4 side chains.  相似文献   

2.
Abstract: A series of potential affinity label derivatives of the amphibian opioid peptide [d ‐Ala2]deltorphin I were prepared by incorporation at the para position of Phe3 (in the ‘message’ sequence) or Phe5 (in the ‘address’ sequence) of an electrophilic group (i.e. isothiocyanate or bromoacetamide). The introduction of the electrophile was accomplished by incorporating Fmoc‐Phe(p‐NHAlloc) into the peptide, followed later in the synthesis by selective deprotection of the Alloc group and modification of the resulting amine. While para substitution decreased the δ‐opioid receptor affinity, selected analogs retained nanomolar affinity for δ receptors. [d ‐Ala2,Phe(p‐NCS)3]deltorphin I exhibited moderate affinity (IC50 = 83 nm ) and high selectivity for δ receptors, while the corresponding amine and bromoacetamide derivatives showed pronounced decreases in δ‐receptor affinity (80‐ and >1200‐fold, respectively, compared with [d ‐Ala2]deltorphin I). In the ‘address’ sequence, the Phe(p‐NH2)5 derivative showed the highest δ‐receptor affinity (IC50 = 32 nm ), while the Phe(p‐NHCOCH2Br)5 and Phe(p‐NCS)5 peptides displayed four‐ and tenfold lower δ‐receptor affinities, respectively. [d ‐Ala2,Phe(p‐NCS)3]deltorphin I exhibited wash‐resistant inhibition of [3H][d ‐Pen2,D‐Pen5]enkephalin (DPDPE) binding to δ receptors at a concentration of 80 nm . [d ‐Ala2, Phe(p‐NCS)3]deltorphin I represents the first affinity label derivative of one of the potent and selective amphibian opioid peptides, and the first electrophilic affinity label derivative of an agonist containing the reactive functionality in the ‘message’ sequence of the peptide.  相似文献   

3.
Abstract: The previously described cyclic mu opioid receptor‐selective tetrapeptide Tyr‐c[d ‐Cys‐Phe‐d ‐Pen]NH2 (Et) (JOM‐6) was modified at residues 1 and 3 by substitution with various natural and synthetic amino acids, and/or by alteration of the cyclic system. Effects on mu and delta opioid receptor binding affinities, and on potencies and efficacies as measured by the [35S]‐GTPγS assay, were evaluated. Affinities at mu and delta receptors were not influenced dramatically by substitution of Tyr1 with conformationally restricted phenolic amino acids. In the [35S]‐GTPγS assay, all of the peptides tested exhibited a maximal response comparable with that of fentanyl at the mu opioid receptor, and all showed high potency, in the range0.4–9 nm . However, potency changes did not always correlate with affinity, suggesting that the conformation required for binding and the conformation required for activation of the opioid receptors are different. At the delta opioid receptor, none of the peptides were able to produce a response equivalent to that of the full delta agonist BW 373,U86 and only one had an EC50 value of less than 100 nm . Lastly, we have identified a peptide, d ‐Hat‐c[d ‐Cys‐Phe‐d ‐Pen]NH2 (Et), with high potency and > 1000‐fold functional selectivity for the mu over delta opioid receptor as measured by the [35S]‐GTPγS assay.  相似文献   

4.
Abstract: A series of cyclic, disulfide‐ or dithioether‐containing tetrapeptides based on previously reported potent μ‐ and δ‐selective analogs has been explored with the aim of improving their poor affinity to the κ‐opioid receptor. Specifically targeted were modifications of tetrapeptide residues 3 and 4, as they presumably interact with residues from transmembrane helices 6 and 7 and extracellular loop 3 that differ among the three receptors. Accordingly, tetrapeptides were synthesized with Phe3 replaced by aliphatic (Gly, Ala, Aib, Cha), basic (Lys, Arg, homo‐Arg), or aromatic sides chains (Trp, Tyr, p‐NH2Phe), and with d ‐Pen4 replaced by d ‐Cys4, and binding affinities to stably expressed μ‐, δ‐, and κ‐receptors were determined. In general, the resulting analogs failed to exhibit appreciable affinity for the κ‐receptor, with the exception of the tetrapeptide Tyr‐c[d ‐Cys‐Phe‐d ‐Cys]‐NH2, cyclized via a disulfide bond, which demonstrated high binding affinity toward all opioid receptors (Kiμ = 1.26 nm , Kiδ = 16.1 nm , Kiκ = 38.7 nm ). Modeling of the κ‐receptor/ligand complex in the active state reveals that the receptor‐binding pocket for residues 3 and 4 of the tetrapeptide ligands is smaller than that in the μ‐receptor and requires, for optimal fit, that the tripeptide cycle of the ligand assume a higher energy conformation. The magnitude of this energy penalty depends on the nature of the fourth residue of the peptide (d ‐Pen or d ‐Cys) and correlates well with the observed κ‐receptor binding affinity.  相似文献   

5.
Abstract: Using results from our previously reported cyclic opioid peptide series and reliable models for μ‐, δ‐, and κ‐opioid receptors (MOR, DOR, and KOR, respectively) and their complexes with peptide ligands, we have designed and synthesized a series of cyclic pentapeptides of structure Tyr‐c[d ‐Cys‐Phe‐Phe‐X]‐NH2, cyclized via disulfide, methylene, or ethylene dithioethers, and where X = d ‐ or l ‐Cys; or d ‐ or l ‐penicillamine (Pen; β,β‐dimethylcysteine). Determination of binding affinities to MOR, DOR, and KOR revealed that members of this series with X = d ‐ or l ‐Cys display KOR affinities in the low nanomolar range, demonstrating that a ‘DPDPE‐like’ tetrapeptide scaffold is suitable not only for DOR and MOR ligands, but also for KOR ligands. The cyclic pentapeptides reported here are not, however, selective for KOR, rather they display significant selectivity and high affinity for MOR. Indeed, peptide 8 , Tyr‐c[d ‐Cys‐Phe‐Phe‐Cys]‐NH2‐cyclized via a methylene dithioether, shows picomolar binding affinity for MOR ( = 16 pm ) with more than 100‐fold selectivity for MOR vs. DOR or KOR, and may be of interest as a high affinity, high selectivity MOR ligand. Nonetheless, the high affinity KOR peptides in this series represent excellent leads for the development of structurally related, selective KOR ligands designed to exploit structurally specific features of KOR, MOR, and DOR.  相似文献   

6.
Abstract: The cyclic enkephalin analog H‐Tyr‐c[d ‐Cys‐Gly‐Phe(pNO2)‐d ‐Cys]NH2 is a highly potent opioid agonist with IC50s of 35 pm and 19 pm in the guinea‐pig ileum (GPI) and mouse vas deferens (MVD) assays, respectively. The Phe1‐analog of this peptide showed 370‐fold and 6790‐fold lower agonist potency in the GPI and MVD assays, respectively, indicating the importance of the Tyr1 hydroxyl‐group in the interaction with μ and δ opioid receptors. In the present study, the effect of various substituents (‐NH2, ‐NO2, ‐CN, ‐CH3, ‐COOH, ‐COCH3, ‐CONH2) introduced in the para‐position of the Phe1‐residue of H‐Phe‐c[d ‐Cys‐Gly‐Phe(pNO2)‐d ‐Cys]NH2 on the in vitro opioid activity profile was examined. Most analogs showed enhanced μ and δ agonist potencies in the two bioassays, except for the Phe(pCOOH)1‐analog, which was weakly active, probably as a consequence of the negative charge. The most potent compounds were the Phe(pCOH3)1‐ and the Phe(pCONH2)1‐analogs. The latter compound showed subnanomolar μ and δ agonist potencies and represents the most potent enkephalin analog lacking the Tyr1 hydroxyl‐group reported to date. Taken together, these results indicate that various substituents introduced in the para‐position of Phe1 enhance opioid activity via hydrogen bonding or hydrophobic interactions with the receptor. Comparison with existing structure‐activity relationship on phenolic hydroxyl replacements in morphinans indicates that these nonpeptide opiates and some of the cyclic enkephalin analogs described here may have different modes of binding to the receptor.  相似文献   

7.
Abstract: Endomorphin‐2 (Tyr‐Pro‐Phe‐Phe‐NH2) binds with high affinity and selectivity to the μ‐opioid receptor. In the present study, [125I]endomorphin‐2 has been used to characterize μ‐opioid‐binding sites on transplantable mouse mammary adenocarcinoma cells. Cold saturation experiments performed with [125I]endomorphin‐2 (1 nm ) show biphasic binding curves in Scatchard coordinates. One component represents high affinity and low capacity (Kd = 18.79 ± 1.13 nm , Bmax = 635 ± 24 fmol/mg protein) and the other shows low affinity and higher capacity (Kd = 7.67 ± 0.81 μm , Bmax = 157 ± 13 pmol/mg protein) binding sites. The rank order of agonists competing for the [125I]endomorphin‐2 binding site was [d ‐1‐Nal3]morphiceptin > endomorphin‐2 ? [d ‐Phe3]morphiceptin > morphiceptin > [d ‐1‐Nal3]endomorphin‐2, indicating binding of these peptides to μ‐opioid receptors. The uptake of 131I‐labeled peptides administered intraperitoneally to tumor‐bearing mice was also investigated. The highest accumulation in the tumor was observed for [d ‐1‐Nal3]morphiceptin, which reached the value of 8.19 ± 1.14% dose/g tissue.  相似文献   

8.
Abstract: We previously reported that the novel dynorphin A (Dyn A, Tyr‐Gly‐Gly‐Phe‐Leu‐Arg‐Arg‐Ile‐Arg‐Pro‐Lys‐Leu‐Lys‐Trp‐Asp‐Asn‐Gln) analog arodyn (Ac[Phe1,2,3,Arg4,d ‐Ala8]Dyn A‐(1–11)NH2, Bennett, M.A., Murray, T.F. & Aldrich, J.V. (2002) J. Med. Chem. vol. 45, pp. 5617–5619) is a κ opioid receptor‐selective peptide [Ki(κ) = 10 nm , Ki ratio (κ/μ/δ) = 1/174/583] which exhibits antagonist activity at κ opioid receptors. In this study, a series of arodyn analogs was prepared and evaluated to explore the structure–activity relationships (SAR) of this peptide; this included an alanine scan of the entire arodyn sequence, sequential isomeric d ‐amino acid substitution in the N‐terminal ‘message’ sequence, NMePhe substitution individually in positions 1–3, and modifications in position 1. The results for the Ala‐substituted derivatives indicated that Arg6 and Arg7 are the most important residues for arodyn's nanomolar binding affinity for κ opioid receptors. Ala substitution of the other basic residues (Arg4, Arg9 and Lys11) resulted in lower decreases in affinity for κ opioid receptors (three‐ to fivefold compared with arodyn). Of particular interest, while [Ala10]arodyn exhibits similar κ opioid receptor binding as arodyn, it displays higher κ vs. μ opioid receptor selectivity [Ki ratio (κ/μ) = 1/350] than arodyn because of a twofold loss in affinity at μ opioid receptors. Surprisingly, the Tyr1 analog exhibits a sevenfold decrease in κ opioid receptor affinity, indicating that arodyn displays significantly different SAR than Dyn A; [Tyr1]arodyn also unexpectedly exhibits inverse agonist activity in the adenylyl cyclase assay using Chinese hamster ovary cells stably expressing κ opioid receptors. Substitution of NMePhe in position 1 gave [NMePhe1]arodyn which exhibits high affinity [Ki(κ) = 4.56 nm ] and exceptional selectivity for κ opioid receptors [Ki ratio (κ/μ/δ) = 1/1100/>2170]. This peptide exhibits antagonistic activity in the adenylyl cyclase assay, reversing the agonism of 10 nm Dyn A‐(1–13)NH2. Thus [NMePhe1]arodyn is a highly κ opioid receptor‐selective antagonist that could be a useful pharmacological tool to study κ opioid receptor‐mediated activities.  相似文献   

9.
Abstract: A series of position 4‐substituted endomorphin‐2 (Tyr‐Pro‐Phe‐Phe‐NH2) analogs containing 3‐(1‐naphthyl)‐alanine (1‐Nal) or 3‐(2‐naphthyl)‐alanine (2‐Nal) in l ‐ or d ‐configuration, was synthesized. The opioid activity profiles of these peptides were determined in the μ‐opioid receptor representative binding assay and in the Guinea‐Pig Ileum assay/Mouse Vas Deferens assay (GPI/MVD) bioassays in vitro, as well as in the mouse hot‐plate test of analgesia in vivo. In the binding assay the affinity of all new analogs for the μ‐opioid receptor was reduced compared with endomorphin‐2. The two most potent analogs were [d ‐1‐Nal4]‐ and [d ‐2‐Nal4]endomorphin‐2, with IC50 values 14 ± 1.25 and 19 ± 2.1 nm , respectively, compared with 1.9 ± 0.21 nm for endomorphin‐2. In the GPI assay these analogs were found to be weak antagonists and they were inactive in the MVD assay. The in vitro GPI assay results were in agreement with those obtained in the in vivo hot‐plate test. Antinociception induced by endomorphin‐2 was reversed by concomitant intracerebroventricula (i.c.v.) administration of [d ‐1‐Nal4]‐ and [d ‐2‐Nal4]‐endomorphin‐2, indicating that these analogs were μ‐opioid antagonists. Their antagonist activity was compared with that of naloxone. At a dose 5 μg per animal naloxone almost completely inhibited antinociceptive action of endomorphin‐2, while [d ‐1‐Nal4]endomorphin‐2 in about 46%.  相似文献   

10.
Abstract: Dynorphin A (Dyn A), a 17 amino acid peptide H‐Tyr‐Gly‐Gly‐Phe‐Leu‐Arg‐Arg‐Ile‐Arg‐Pro‐Lys‐Leu‐Lys‐Trp‐Asp‐Asn‐Gln‐OH, is a potent opioid peptide which interacts preferentially with κ‐opioid receptors. Research in the development of selective and potent opioid peptide ligands for the κ‐receptor is important in mediating analgesia. Several cyclic disulphide bridge‐containing peptide analogues of Dyn A, which were conformationally constrained in the putative message or address segment of the opioid ligand, were designed, synthesized and assayed. To further investigate the conformational and topographical requirements for the residues in positions 5 and 11 of these analogues, a systematic series of Dyn A1?11‐NH2 cyclic analogues incorporating the sulphydryl‐containing amino acids l ‐ and d ‐Cys and l ‐ and d ‐Pen in positions 5 and 11 were synthesized and assayed. Cyclic lactam peptide analogues were also synthesized and assayed. Several of these cyclic analogues, retained the same affinity and selectivity (vs. the μ‐ and δ‐receptors) as the parent Dyn A1?11‐NH2 peptide in the guinea‐pig brain (GPB), but exhibited a much lower activity in the guinea‐pig ileum (GPI), thus leading to centrally vs. peripherally selective peptides. Studies of the structure–activity relationship of Dyn A peptide provide new insights into the importance of each amino acid residue (and their configurations) in Dyn A analogues for high potency and good selectivity at κ‐opioid receptors. We report herein the progress towards the development of Dyn A peptide ligands, which can act as agonists or antagonists at cell surface receptors that modulate cell function and animal behaviour using various approaches to rational peptide ligand‐based drug design.  相似文献   

11.
Abstract: Dermorphin and [Lys7]dermorphin, selective µ‐opioid receptor ligands originating from amphibian skin, have been modified with various electrophiles in either the ‘message’ or ‘address’ sequences as potential peptide‐based affinity labels for µ‐receptors. Introduction of the electrophilic isothiocyanate and bromoacetamide groups on the para position of Phe3 and Phe5 was accomplished by incorporating Fmoc‐Phe(p‐NHAlloc) into the peptide followed by selective deprotection and modification. The corresponding amine‐containing peptides were also prepared. The pure peptides were evaluated in radioligand binding experiments using Chinese hamster ovary (CHO) cells expressing µ‐ and δ‐opioid receptors. In dermorphin, introduction of the electrophilic groups in the ‘message’ domain lowered the binding affinity by > 1000‐fold; only [Phe(p‐NH2)3]dermorphin retained nanomolar affinity for µ‐receptors. Modifications in the ‘address’ region of both dermorphin and [Lys7]dermorphin were relatively well tolerated. In particular, [Phe(p‐NH2)5,Lys7]dermorphin showed similar affinity to dermorphin, with almost 2‐fold higher selectivity for µ‐receptors. [Phe(p‐NHCOCH2Br)5]‐ and [Phe(p‐NHCOCH2Br)5,Lys7]dermorphin exhibited relatively high affinity (IC50 = 27.7 and 15.1 nm , respectively) for µ‐receptors. However, neither of these peptides inhibited [3H]DAMGO binding in a wash‐resistant manner.  相似文献   

12.
Abstract: In an attempt to identify potential peptide‐based affinity labels for opioid receptors, endomorphin‐2 (Tyr‐Pro‐Phe‐PheNH2), a potent and selective endogenous ligand for µ‐opioid receptors, was chosen as the parent peptide for modification. The tetrapeptide analogs were prepared using standard Fmoc‐solid phase peptide synthesis in conjunction with incorporation of Fmoc‐Phe(p‐NHAlloc) and modification of the p‐amino group. The electrophilic groups isothiocyanate and bromoacetamide were introduced into the para position on either Phe3 or Phe4; the corresponding free amine‐containing peptides were also prepared for comparison. The peptides bearing an affinity label group and their free amine analogs were evaluated in a radioligand‐binding assay using Chinese hamster ovary (CHO) cells expressing µ‐ and δ‐opioid receptors. Modification on Phe4 was better tolerated than on Phe3 for µ‐receptor binding. Among the analogs tested, [Phe(p‐NH2)4]endomorphin‐2 showed the highest affinity (IC50 = 37 nm ) for µ‐receptors. The Phe(p‐NHCOCH2Br)4 analog displayed the highest µ‐receptor affinity (IC50 = 158 nm ) among the peptides containing an affinity label group. Most of the compounds exhibited negligible binding affinity for δ‐receptors, similar to the parent peptide.  相似文献   

13.
Abstract: In order to extend the use of proteases to organic synthesis and seek the rules of enzymatic reactions in organic media, we focused on unnatural substrates for proteases to form amide bonds. In this paper, the study of unnatural substrates containing d ‐amino acid residue, which act as acyl acceptors as well as acyl donors for proteases in organic media, is reported. Dermorphin is a heptapeptide (H‐Tyr‐d ‐Ala‐Phe‐Gly‐Tyr‐Pro‐Ser‐NH2) with potent analgesic activity. The N‐terminal tetrapeptide is the minimum sequence that retains dermorphin activity, and is selected as the model compound in our study. Two dermorphin‐(1–4) derivatives, Boc‐Tyr‐d ‐Ala‐Phe‐Gly‐N2H2Ph and Boc‐Tyr‐d ‐Ala‐Phe‐Gly‐NH2, which contained a d ‐amino acid residue, were synthesized by proteases in organic media for the first time. The synthesis of these two dermorphin‐(1–4) derivatives could be catalyzed by subtilisin with Boc‐Tyr‐d ‐Ala‐OCH2CF3 as an acyl donor substrate in AcOEt. The synthesis of dermorphin‐(1–2) derivative Boc‐Tyr‐d ‐Ala‐N2H2Ph was catalyzed by α‐chymotrypsin in different organic solvents and d ‐Ala‐N2H2Ph was used as an acyl acceptor substrate. Factors influencing the above enzymatic reactions were systematically studied.  相似文献   

14.
Abstract: Solid‐phase synthetic methodology was developed for the preparation of peptide‐based affinity labels. The initial peptides synthesized were dynorphin A (Dyn A) analogs [Phe(p‐X)4,d ‐Pro10]Dyn A(1–11)NH2 containing isothiocyanate (X = –N=C=S) and bromoacetamide (X = –NHCOCH2Br) groups. The peptides were assembled on solid supports using Fmoc‐protected amino acids, and the side chain amine to be functionalized, Phe(p‐NH2), was protected by the Alloc (allyloxycarbonyl) group. Following removal of the Alloc group by palladium(0), the reactive isothiocyanate and bromoacetamide functionalities were successfully introduced while the peptides were still attached to the resin. Synthesis of these peptides was carried out on polystyrene (PS) and polyethylene glycol–polystyrene (PEG–PS) resins containing the PAL [peptide amide linker, 5‐(4‐Fmoc‐aminomethyl‐3,5‐dimethoxyphenoxy)valeric acid] linker. Both the rate of Alloc deprotection and the purity of the crude affinity‐labeled peptides obtained were found to be dependent on the resin used for peptide assembly.  相似文献   

15.
Abstract: There is evidence to indicate that opioid compounds with mixed μ agonist/δ antagonist properties are analgesics with low propensity to produce tolerance and physical dependence. A chimeric peptide containing the potent and selective μ agonist H‐Dmt‐D‐Arg‐Phe‐Lys‐NH2 ([Dmt1]DALDA) (Dmt = 2′,6′‐dimethyltyrosine) and the potent and selective δ antagonist H‐Tyr‐TicΨ[CH2‐NH]Cha‐Phe‐OH (TICP[Ψ]) (Cha = cyclohexylalanine), connected ‘tail‐to‐tail’ via a short linker, was synthesized using a combination of solid‐phase and solution techniques. The resulting peptide, H‐Dmt→D‐Arg→Phe→Lys‐NH‐CH2‐CH2‐NH‐Phe←Cha[NH‐CH2]ΨTic←Tyr‐H, showed the expected μ agonist/δ antagonist profile in the guinea‐pig ileum and mouse vas deferens assays. Its μ and δ receptor binding affinities were in the low nanomolar range, as determined in rat brain membrane binding assays.  相似文献   

16.
Abstract: The synthesis of conformationally restricted dipeptidic moieties 4‐amino‐1,2,4,5‐tetrahydro‐2‐benzazepin‐3‐one (Aba)‐Gly ([(4S)‐amino‐3‐oxo‐1,2,4,5‐tetrahydro‐1H‐2‐benzazepin‐2‐yl]‐acetic acid) and 8‐hydroxy‐4‐amino‐1,2,4,5‐tetrahydro‐2‐benzazepin‐3‐one (Hba)‐d ‐Ala ([(4S)‐amino‐8‐hydroxy‐3‐oxo‐1,2,4,5‐tetrahydro‐benzo[c]azepin‐2‐yl]‐propionic acid) was based on a synthetic strategy that uses an oxazolidinone as an N‐acyliminium precursor. Introducing these Aba scaffolds into the N‐terminal tetrapeptide of dermorphin (H‐Tyr‐d ‐Ala‐Phe‐Gly‐Tyr‐Pro‐Ser‐NH2)‐induced remarkable shifts in affinity and selectivity towards the opioid μ‐ and δ‐receptors. This paper provides the synthesis and biological in vitro and in vivo evaluation of constricted analogues of the N‐terminal tetrapeptide H‐Tyr‐d ‐Ala‐Phe‐Gly‐NH2, which is the minimal subunit of dermorphin needed for dermorphin‐like opiate activity.  相似文献   

17.
This article describes new deltorphin I analogs in which phenylalanine residues were replaced by the corresponding (R) or (S)‐α‐benzyl‐β‐azidoalanine, α‐benzyl‐β‐(1‐pyrrolidinyl)alanine, α‐benzyl‐β‐(1‐piperidinyl)alanine, and α‐benzyl‐β‐(4‐morpholinyl)‐alanine residues. The potency and selectivity of the new analogs were evaluated by a competitive receptor binding assay in the rat brain using [3H]DAMGO (a μ ligand) and [3H]DELT (a δ ligand). The affinity of analogs containing (R) or (S)‐α‐benzyl‐β‐azidoalanine in position 3 to δ‐receptors strongly depended on the chirality of the α,α‐disubstituted residue. The conformational behavior of peptides modified with (R) or (S)‐α‐benzyl‐β‐(1‐piperidinyl)Ala, which displays the opposite selectivity, was analyzed by 1H and 13C NMR. The μ‐selective Tyr‐d ‐Ala‐(R)‐α‐benzyl‐β‐(1‐piperidinyl)Ala‐Asp‐Val‐Val‐Gly‐NH2 lacks the helical conformation observed in the δ‐selective Tyr‐d ‐Ala‐(S)‐α‐benzyl‐β‐(1‐piperidinyl)Ala‐Asp‐Val‐Val‐Gly‐NH2. Our results support the proposal that differences between δ‐ and μ‐selective opioid peptides are attributable to the presence or absence of a spatial overlap between the N‐terminal message domain and the C‐terminal address domain.  相似文献   

18.
We report the synthesis and binding properties of specific photoaffinity ligands for μ and δ opioid receptor subtypes. These ligands are derived from DAGO: Tyr-D-Ala-Gly-NMePhe-Gly-ol, a μ selective probe and DTLET: Tyr-D-Thr-Gly-Phe-Leu-Thr, a δ selective probe by modifying the Phe 4 residue. These modifications are: i) a nitro group on the para position of Phe ring as Phe(4 NO2) or Nip, ii) an azido group as Phe(4 N3) or AZ. Pharmacological responses on mouse vas deferens (δ sites) and guinea pig ileum (μ sites), as well as competition experiments with [3H] DAGO and [3H] DTLET on crude rat brain membranes have been performed. The nitro group on the phenyl ring of the Phe residue preserves the affinity and selectivity of each probe: NipDAGO for the μ sites, NipDTLET for the δ ones. However the nitro probes do not appear to be photo-activable by u.v. irradiation. Likewise, azidation of the phenyl ring of the Phe residue does not change the receptor selectivity of each probe, but AZDAGO has less affinity than its parent molecule DAGO, while AZDTLET has more affinity than DTLET. These compounds are photoactivable and provide an efficient tool to characterize and isolate the different receptor subtypes, especially the δ site.  相似文献   

19.
Abstract: Replacement of Phe3 in the endogenous δ‐opioid selective peptide deltorphin I with four optically pure stereoisomers of the topographically constrained, highly hydrophobic novel amino acid β‐isopropylphenylalanine (β‐iPrPhe) produced four pharmacologically different deltorphin I peptidomimetics. Radiolabeled ligand‐binding assays and in vitro biological evaluation indicate that the stereoconfiguration of the iPrPhe residue plays a crucial role in determining the binding affinity, bioactivity and selectivity of [β‐iPrPhe3]deltorphin I analogs: a (2S,3R) configuration of the iPrPhe3 residue in [β‐iPrPhe3]deltorphin I provided the most desirable biological properties with binding affinity (IC50 = 2 n m ), bioassay potency (IC50 = 1.23 n m in MVD assay) and exceptional selectivity for the δ‐opioid receptor over the µ‐opioid receptor (30 000). Further conformational studies based on two‐dimensional NMR and computer‐assisted molecular modeling suggested a model for the possible bioactive conformation in which the Tyr1 and (2S,3R)‐β‐iPrPhe3 residues adopt trans side‐chain conformations, and the linear peptide backbone favors a distorted β‐turn conformation.  相似文献   

20.
Abstract: The dermorphin‐derived tetrapeptide H‐Dmt‐d ‐Arg‐Phe‐Lys‐NH2 (Dmt = 2′,6′‐dimethyltyrosine) ([Dmt1]DALDA) is a highly potent and selective μ‐opioid agonist capable of crossing the blood–brain barrier and producing a potent, centrally mediated analgesic effect when given systemically. For the purpose of biodistribution studies by fluorescence techniques, [Dmt1]DALDA analogues containing various fluorescent labels [dansyl, anthraniloyl (atn), fluorescein, or 6‐dimethylamino‐2′‐naphthoyl] in several different locations of the peptide were synthesized and characterized in vitro in the guinea‐pig ileum and mouse vas deferens assays, and in μ‐, δ‐ and κ‐opioid receptor‐binding assays. The analogues showed various degrees of μ receptor‐binding selectivity, but all of them were less μ‐selective than the [Dmt1]DALDA parent peptide. Most analogues retained potent, full μ‐agonist activity, except for one with fluorescein attached at the C‐terminus ( 3a ) (partial μ‐agonist) and one containing β‐(6′‐dimethylamino‐2′‐naphthoyl)alanine (aladan) in place of Phe3 ( 4 ) (μ‐ and κ‐antagonist). The obtained data indicate that the receptor‐binding affinity, receptor selectivity and intrinsic efficacy of the prepared analogues vary very significantly, depending on the type of fluorescent label used and on its location in the peptide. The results suggest that the biological activity profile of fluorescence‐labeled peptide analogues should always be carefully determined prior to their use in biodistribution studies or other studies. One of the analogues containing the atn group ( 2a ) proved highly useful in a study of cellular uptake and intracellular distribution by confocal laser scanning microscopy.  相似文献   

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