RESUMO
A comparative molecular modeling study of delta-opioid ligands was performed under the assumption that potent peptide and nonpeptide agonists may have common three-dimensional (3D) arrangement of pharmacophore groups upon binding to the delta-receptor. Low-energy conformations of the agonists 7-spiroindanyloxymorphone (SIOM) and 2-methyl-4a-alpha-(3-hydroxyphenyl)-1,2,3,4,4a,5,12, 12a-alpha-octahydro-quinolino[2,3,3-g]isoquinoline (TAN-67), and a partial agonist oxomorphindole (OMI) were determined by high-temperature molecular dynamics (MD). A good spatial overlap was found for the pharmacophore groups of SIOM, TAN-67, and OMI, including the basic nitrogen, phenol hydroxyl, and two aromatic ring. Based on this overlap we proposed a 3D pharmacophore model for nonpeptide delta-opioid agonists with a distance of 7.0 +/- 1.3 A between the two aromatic rings and of 8.2 +/- 1.0 A between the nitrogen and phenyl ring. The potent and highly delta-opioid receptor selective agonist [(2S,3R)-TMT(1)]DPDPE, which shares global backbone constraints of the 14-membered disulfide cycle and a strong preference for the trans rotamer of the TMT(1) side chain, was chosen as a peptide template of the delta-opioid pharmacophore. Extensive MD simulations at 300 K with the AMBER force field were performed for [(2S,3R)-TMT(1)]DPDPE and the less potent [(2S, 3S)-TMT(1)]DPDPE analogue. Multiple MD trajectories were collected for each peptide starting from the x-ray structures of DPDPE and [L-Ala(3)]DPDPE and from models proposed in the literature. Low-energy MD conformations were filtered by the nonpeptide pharmacophore query and then directly superimposed with SIOM, OMI, and TAN-67. Two conformers of [(2S,3R)-TMT(1)]DPDPE that showed the best overlap with the nonpeptide pharmacophore (rms deviation = 1. 0 A for N,O atoms and centroids of two aromatic rings) were selected as possible delta-receptor binding conformations. These conformations have similar backbone structures, and trans rotamers of the TMT(1) side-chain group. They are reasonably close to the crystal structure of [L-Ala(3)]DPDPE, and differ significantly from the crystal structure of DPDPE. The conformer with a gauche(-) rotamer of Phe(4) is most consistent with structure-activity relationships of delta-opioid peptides. The proposed 3D models were used for rational design of new nonpeptide delta-receptor ligands.
Assuntos
D-Penicilina (2,5)-Encefalina/química , Ligantes , Oximorfona/análogos & derivados , Peptídeos/química , Quinolinas/química , Receptores Opioides delta/química , Compostos de Espiro/química , Analgésicos/química , Sítios de Ligação , D-Penicilina (2,5)-Encefalina/análogos & derivados , Modelos Moleculares , Conformação Molecular , Morfolinas/química , Oximorfona/química , Relação Estrutura-AtividadeRESUMO
On the basis of the structure-activity relationships of delta-opioid-selective peptide ligands and on a model of the proposed bioactive conformation for a potent and selective, conformationally constrained delta-opioid peptide ligand [(2S, 3R)-TMT1]DPDPE, a series of small organic peptide mimetic compounds targeted for the delta-opioid receptor have been designed, synthesized, and evaluated in radiolabeled ligand binding assays and in vitro bioassays. The new non-peptide ligands use piperazine as a template to present the most important pharmacophore groups, including phenol and phenyl groups and a hydrophobic moiety. This hydrophobic group was designed to mimic the hydrophobic character of the D-Pen residues in DPDPE, which has been found to be extremely important for increasing the binding affinity and selectivity of these non-peptide ligands for the delta-opioid receptor over the mu-opioid receptor. Compound 6f (SL-3111) showed 8 nM binding affinity and over 2000-fold selectivity for the delta-opioid receptor over the mu-opioid receptor. Both enantiomers of SL-3111 were separated, and the (-)-isomer was shown to be the compound with the highest affinity for the delta-opioid receptor found in our study (IC50 = 4.1 nM), with a selectivity very similar to that observed for the racemic compound. The phenol hydroxyl group of SL-3111 turned out to be essential to maintain high affinity for the delta-opioid receptor, which also was observed in the case of the delta-opioid-selective peptide ligand DPDPE. Binding studies of SL-3111 and [p-ClPhe4]DPDPE on the cloned wild-type and mutated human delta-opioid receptors suggested that the new non-peptide ligand has a binding profile similar to that of DPDPE but different from that of (+)-4-[((alphaR)-alpha(2S,5R)-4-allyl-2, 5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC-80), another delta-opioid-selective non-peptide ligand.
Assuntos
Compostos Benzidrílicos/síntese química , Peptídeos/química , Piperazinas/síntese química , Receptores Opioides delta/agonistas , Animais , Compostos Benzidrílicos/química , Compostos Benzidrílicos/metabolismo , Compostos Benzidrílicos/farmacologia , Encéfalo/metabolismo , Desenho de Fármacos , Cobaias , Humanos , Íleo/efeitos dos fármacos , Íleo/fisiologia , Técnicas In Vitro , Ligantes , Masculino , Camundongos , Camundongos Endogâmicos ICR , Mimetismo Molecular , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Músculo Liso/fisiologia , Mutação , Piperazinas/química , Piperazinas/metabolismo , Piperazinas/farmacologia , Ensaio Radioligante , Ratos , Receptores Opioides delta/genética , Receptores Opioides delta/metabolismo , Estereoisomerismo , Relação Estrutura-Atividade , Ducto Deferente/efeitos dos fármacos , Ducto Deferente/fisiologiaRESUMO
Extensive evidence gathered from structure-activity relationship analysis has identified and confirmed specific positions in the glucagon sequence that are important either for binding to its receptor or for signal transduction. Fifteen glucagon analogues have been designed and synthesized by incorporating structural changes in the N-terminal region of glucagon, in particular histidine-1, phenylalanine-6, and aspartic acid-9. This investigation was conducted to study the role of phenylalanine at position 6 on the glucagon mechanism of action. These glucagon analogues have been made by either deleting or substituting hydrophobic groups, hydrophilic groups, aromatic amino acids, or a D-phenylalanine residue at this position. The structures of the new analogues are as follows: [des-His1, des-Phe6, Glu9]glucagon-NH2 (1); [des-His1,Ala6,Glu9]glucagon-NH2 (2); [des-His1,Tyr6,Glu9]glucagon-NH2 (3); [des-His1,Trp6,Glu9]-glucagon-NH2 (4); [des-His1,D-Phe6,Glu9]glucagon-NH2 (5); [des-His1,Nle6,Glu9]glucagon-NH2 (6); [des-His1,Asp6,Glu9]glucagon-NH2 (7); [des-His1,des-Gly4,Glu9]glucagon-NH2 (8); [desPhe6,-Glu9]glucagon-NH2 (9); [des-Phe6]glucagon-NH2 (10); [des-His1, des-Phe6]glucagon-NH2 (11); [des-His1, des-Phe6,Glu9]glucagon (12); [des-Phe6,Glu9]glucagon (13); [des-Phe6]glucagon (14); and [des-His1, des-Phe6]glucagon (15). The receptor binding potencies IC50 values are 48 (1), 126 (2), 40 (3), 19 (4), 100 (5), 48 (6), 2000 (7), 52 (8), 113 (9), 512 (10), 128 (11), 1000 (12), 2000 (13), 500 (14), and 200 nM (15). All analogues were found to be antagonists unable to activate the adenylate cyclase system even at concentrations as high as 10(-5) M except for analogues 6 and 8, which were found to be weak partial agonists/partial antagonists with maximum stimulation between 6-12%. In competitive inhibition experiments, all the analogues caused a right shift of the glucagon-stimulated adenylate cyclase dose-response curve. The pA2 values were 8.20 (1), 6.40 (2), 6.20 (3), 6.25 (4), 6.30 (5), 6.30 (7), 6.05 (8), 6.20 (9), 6.30 (10), 6.25 (11), 6.10 (12), 6.20 (13), 6.20 (14), and 6.35 (15).
Assuntos
Glucagon/metabolismo , Fenilalanina/metabolismo , Receptores de Glucagon/metabolismo , Adenilil Ciclases/metabolismo , Sequência de Aminoácidos , Animais , Membrana Celular/metabolismo , Cromatografia Líquida de Alta Pressão , Cristalografia por Raios X , Glucagon/análogos & derivados , Fígado/metabolismo , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Dados de Sequência Molecular , Ratos , Ratos Sprague-Dawley , Receptores de Glucagon/antagonistas & inibidores , Relação Estrutura-AtividadeRESUMO
The role of position 10 in the beta-turn region of glucagon was investigated by substituting chiral constrained amino acids and other modifications in the N-terminal region. A series of glucagon analogues have been designed and synthesized by incorporating beta-methylphenylalanine isomers (2S,3S, 2S,3R, 2R,3R, and 2R,3S) at position 10 in order to explore the structural and topographical requirements of the glucagon receptor, and, in addition, utilizing previous studies which indicated that antagonism could be enhanced by modifications (des-His1, Glu9) and a bulky group at position 5. The structures of the new analogues are as follows: [des-His1,-Tyr5,Glu9]glucagon-NH2 (II), [des-His1,Tyr5,Glu9,Phe10]glucagon-NH2 (III), [des-His1,Tyr5,Glu9,-Ala10]glucagon-NH2 (IV), [des-His1,Tyr5,Glu9,(2S,3R)-beta-MePhe10]glucagon-NH2 (V), [des-His1,-Tyr5,Glu9,(2S,3S)-beta-MePhe10]glucagon-NH2 (VI), [des-His1,Tyr5,Glu9,D-Tyr10]glucagon-NH2 (VII), [des-His1,Tyr5,Glu9,D-Phe10]glucagon-NH2 (VIII), [des-His1,Tyr5,Glu9,D-Ala10]glucagon-NH2 (IX), [des-His1,Tyr5,Glu9,(2R,3R)-beta-MePhe10]glucagon-NH2 (X), and [des-His1,Tyr5,Glu9,(2R,3S)-beta-MePhe10]glucagon-NH2 (XI). These analogues led to dramatically different changes in in vitro binding affinities for glucagon receptors. Their receptor binding potencies IC50 values (nM) are 2.3 (II), 4.1 (III), 395.0 (IV), 10.0 (V), 170.0 (VI), 74.0 (VII), 34.5 (VIII), 510.0 (IX), 120.0 (X), and 180.0 (XI). Analogues II, III, V, VI, and XI were found to be weak partial agonists/partial antagonists with maximum stimulation between 5%-9%, while the other compounds (IV and VII-X) were antagonists unable to activate the adenylate cyclase system even at concentrations as high as 10(-5) M. In competition experiments, all of the analogues caused a right shift of the glucagon-stimulated adenylate cyclase dose-response curve. The pA2 values were 6.60 (II), 6.85 (III), 6.20 (IV), 6.20 (V), 6.10 (VI), 6.50 (VII), 6.20 (VIII), 5.85 (IX), 6.20 (X), and 6.00 (XI). Putative topographical requirements of the glucagon receptor for the aromatic side chain conformation in position 10 of glucagon antagonists are discussed.
Assuntos
Glucagon/análogos & derivados , Glucagon/metabolismo , Peptídeos/síntese química , Receptores de Glucagon/metabolismo , Adenilil Ciclases/metabolismo , Sequência de Aminoácidos , Aminobutiratos/química , Animais , Membrana Celular/metabolismo , Glucagon/química , Glucagon/farmacologia , Fígado/enzimologia , Dados de Sequência Molecular , Peptídeos/metabolismo , Peptídeos/farmacologia , Ligação Proteica , Conformação Proteica , Estrutura Secundária de Proteína , Ratos , Ratos Sprague-Dawley , Receptores de Glucagon/agonistas , Receptores de Glucagon/antagonistas & inibidores , Transdução de Sinais , Relação Estrutura-AtividadeRESUMO
Solution conformations of beta-methyl-para-nitrophenylalanine4 analogues of the potent delta-opioid peptide cyclo[D-Pen2, D-Pen5]enkephalin (DPDPE) were studied by combined use of nmr and conformational energy calculations. Nuclear Overhauser effect connectivities and 3JHNC alpha H coupling constants measured for the (2S, 3S)-, (2S, 3R)-, and (2R, 3R)-stereoisomers of [beta-Me-p-NO2Phe4]DPDPE in DMSO were compared with low energy conformers obtained by energy minimization in the Empirical Conformational Energy Program for Peptides (ECEPP/2) force field. The conformers that satisfied all available nmr data were selected as probable solution conformations of these peptides. Side-chain rotamer populations, established using homonuclear (3JH alpha H beta) and heteronuclear (3JH alpha C gamma) coupling constants and 13C chemical shifts, show that the beta-methyl substituent eliminates one of the three staggered rotamers of the torsion angle chi 1 for each stereoisomer of the beta-Me-p-NO2Phe4. Similar solution conformations were suggested for the L-Phe4-containing (2S, 3S)- and (2S, 3R)-stereoisomers. Despite some local differences, solution conformations of L- and D-Phe4-containing analogues have a common shape of the peptide backbone and allow similar orientations of the main delta-opioid pharmacophores. This type of structure differs from several models of the solution conformations of DPDPE, and from the model of biologically active conformations of DPDPE suggested earlier. The latter model is allowed for the potent (2S, 3S)- and (2S, 3R)-stereoisomers of [beta-Me-p-NO2Phe4]DPDPE, but it is forbidden for the less active (2R, 3R)- and (2R, 3S)-stereoisomers. It was concluded that the biologically active stereoisomers of [beta-Me-p-NO2Phe4]DPDPE in the delta-receptor-bound state may assume a conformation different from their favorable conformations in DMSO.
Assuntos
Encefalinas/metabolismo , Sequência de Aminoácidos , D-Penicilina (2,5)-Encefalina , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Molecular , Conformação Proteica , Estereoisomerismo , TermodinâmicaRESUMO
Conformational analysis of the neurohypophyseal hormones oxytocin (OT) and arginine-vasopressin (AVP) has been carried out using two different computational approaches and three force fields, namely by the Electrostatically Driven Monte Carlo (EDMC) method, with the Empirical Conformational Energy Program for Peptides (ECEPP/3) force field or with the ECEPP/3 force field plus a hydration-shell model, and by simulated-annealing molecular dynamics with the Consistent Valence Force Field (CVFF). The low-energy conformations obtained for both hormones were classified using the minimal-tree clustering algorithm and characterized according to the locations of beta-turns in the cyclic moieties. Calculations with the CVFF force field located conformations with a beta-turn at residues 3 and 4 as the lowest energy ones both for OT and for AVP. In the ECEPP/3 force field the lowest energy conformation of OT contained a beta-turn at residues 2 and 3, conformations with this location of the turn being higher in energy for AVP. The latter difference can be attributed to the difference in the size of the side chain in position 3 of the sequences: the bulkier phenylalanine residue of AVP in combination with the bulky Tyr2 residue hinders the formation of a turn at residues 2 and 3. Conformations of OT and AVP with a turn at residues 3,4 were in the best agreement with the x-ray structures of deaminooxytocin and pressinoic acid (the cyclic moiety of vasopressin), respectively, and with the nmr-derived distance constraints. Generally, the low-energy conformations obtained with the hydration-shell model were in a better agreement with the experimental data than the conformations calculated in vacuo. It was found, however, that the obtained low-energy conformations do not satisfy all of the nmr-derived distance constraints and the nuclear Overhauser effect pattern observed in nmr studies can be fully explained only by assuming a dynamic equilibrium between conformations with beta-turns at residues 2,3, 3,4, and 4,5. The low-energy structures of OT with a beta-turn at residues 2,3 have the disulfide ring conformations close to the model proposed recently for a potent bicyclic antagonist of OT [M. D. Shenderovich et al. (1994) Polish Journal of Chemistry, Vol. 25, pp. 921-927], although the native hormone differs from the bicyclic analogue by the conformation of the C-terminal tripeptide. This finding confirms the hypothesis of different receptor-bound conformations of agonists and antagonists of OT.
Assuntos
Arginina Vasopressina/química , Ocitocina/química , Sequência de Aminoácidos , Cristalografia por Raios X , Eletroquímica , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Dados de Sequência Molecular , Método de Monte Carlo , Conformação Proteica , TermodinâmicaRESUMO
Seven side chain-constrained bicyclic alpha-melanotropin (alpha-MSH) analogues were designed and synthesized, their conformations analyzed, and their biological properties examined in the frog skin and lizard skin bioassays. The structure of these analogues is based on the central sequence Ac-Cys4-Xaa5-His6-DPhe7-Arg8-Trp9-Cys10-Lys11 -NH2 (Xaa5 = Asp or Glu) and has been extended on the N-terminal with the amino acids Ser1-Tyr2-Ser3 and on the C-terminal with Pro12-Val13 to more closely resemble the native hormone alpha-MSH. The analogue Ac-Cys4-Asp5-His6-DPhe7-Arg8-Trp9-Lys10-Cys11 -NH2 also was synthesized, and its conformational and biological properties were examined. Design of these analogues was based upon the previously identified superpotent monocyclic peptides [Cys4,DPhe7,Cys10]alpha-MSH(4-10)-NH2 and [Nle4,Asp5,DPhe7,Lys10]alpha-MSH(4-10)-NH2 with the rationale of increasing conformational constraints to restrict the available backbone conformations as a means to identify the conformations that facilitate biological activity. Computer-assisted conformational analysis of the central tetrapeptide residues 6-9 identified beta-turns which varied with respect to the residue in the i + 1 position. Each highly constrained peptide contains D-Phe7 and a 23-membered ring which has previously been identified as crucial to produce prolonged acting peptides with superagonistic activities. The bicyclic peptides reported in this study are full agonists and are 25-400-fold less potent than alpha-MSH in the frog and lizard skin bioassays.
Assuntos
Desenho de Fármacos , alfa-MSH/síntese química , Sequência de Aminoácidos , Animais , Bioensaio , Lagartos , Dados de Sequência Molecular , Conformação Proteica , Rana pipiens , Pele/citologia , Pele/efeitos dos fármacos , alfa-MSH/química , alfa-MSH/farmacologiaRESUMO
Conformational energy calculations were carried out on three non-peptide antagonists of oxytocin and vasopressin: penicilide (compound 1; selective for oxytocin receptors), 1-¿1-[4-(3-acetylaminopropoxy(benzoyl]-4-piperidyl¿-3,4-dihydro-2( 1H)-quinoline (compound 2; selective for vasopressin V1 receptors) and 5-dimethylamino-1-¿(2-methylbenzylamino)-benzoyl¿-2,3,4,5-tetrahyd ro-1H-benzapine (compound 3; selective for vasopressin V2 receptors). The obtained low-energy conformations of compound 1 were compared with low-energy conformations of oxytocin (OT) and low-energy conformations of compounds 2 and 3 were compared with low-energy conformations of arginine vasopressin (AVP). It was found that the affinity of the non-peptide antagonists and their selectivity for vasopressin and oxytocin receptors is probably connected with mimicking the aromatic rings of the Tyr2 and the Phe3 residues of AVP in the case of compounds 2 and 3 and with mimicking the Tyr2 residue and the Ile3 or Leu8 residues of OT by the outer benzene ring and the isobutyl group of compound 1. Application of the results in the design of more potent non-peptide antagonists of OT and VP is also discussed.
Assuntos
Receptores de Ocitocina/metabolismo , Receptores de Vasopressinas/metabolismo , Antagonistas dos Receptores de Hormônios Antidiuréticos , Benzazepinas/química , Benzazepinas/metabolismo , Benzazepinas/farmacologia , Técnicas In Vitro , Cinética , Lactonas/química , Lactonas/metabolismo , Lactonas/farmacologia , Modelos Moleculares , Conformação Molecular , Ocitocina/química , Piperidinas/química , Piperidinas/metabolismo , Piperidinas/farmacologia , Quinolonas/química , Quinolonas/metabolismo , Quinolonas/farmacologia , Receptores de Ocitocina/antagonistas & inibidores , Termodinâmica , Vasopressinas/químicaRESUMO
Topographic design of peptide ligands using specialized topographically constrained amino acids can provide new insights into the stereochemical requirements for delta opioid receptors. A highly constrained tyrosine derivative, (2S,3S)-beta-methyl-2',6'-dimethyltyrosine [(2S,3S)-TMT], was prepared by asymmetric synthesis and incorporated in [D-Pen2,D-Pen5] enkephalin (delta 1) and Deltorphin I (delta 2). The results of binding assays and bioassays showed that the two analogues (3 and 4) acted very differently at delta opioid receptors. Further pharmacological evaluations suggested that they actually interact primarily with the delta 1 and delta 2 receptor subtypes, respectively. These results, and conformational studies using NMR and computer-assisted modeling, provided insights into the different stereochemical requirements for these two delta opioid ligands to recognize the delta opioid receptor and its subtypes.
Assuntos
Analgésicos/química , Metiltirosinas/química , Peptídeos/química , Receptores Opioides delta/efeitos dos fármacos , Sequência de Aminoácidos , Analgésicos/metabolismo , Analgésicos/farmacologia , Animais , D-Penicilina (2,5)-Encefalina , Encefalinas/química , Cobaias , Técnicas In Vitro , Espectroscopia de Ressonância Magnética , Masculino , Metiltirosinas/metabolismo , Metiltirosinas/farmacologia , Camundongos , Camundongos Endogâmicos ICR , Dados de Sequência Molecular , Oligopeptídeos/química , Peptídeos/metabolismo , Peptídeos/farmacologia , Conformação Proteica , Ratos , Ratos Sprague-Dawley , Receptores Opioides delta/metabolismo , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
To compare features of the receptor-binding sites (RBSs) of different influenza virus hemagglutinins (HA), binding of a number of synthetic sialic acid (SA) analogs and natural sialosides by a panel of about 30 human influenza A and B virus strains was studied in a competitive ligand binding assay. For all the viruses tested, the N-acetyl group of Neu5Ac, as well as the natural orientation of the carboxylic group at C2 and the hydroxylic group at C4, was essential for binding. Significant type- and subtype-specific differences were observed in virus recognition of asialic parts of sialosides. H1 strains, unlike H3 and type B viruses, were found to bind alpha 2-6-sialyl-N-acetyllactosamine with about an order of magnitude higher affinity than alpha 2-6-sialyllactose (6'SL). The H1 viruses and the H3 strains with Gln in position 226 of HA, but not the H3 strains with Leu-226, bound 6'SL with a lower affinity than alpha 2-3-sialyllactose; this effect correlated clearly with the preferential binding by the former viruses of unsubstituted alpha Neu5Ac compared to methyl alpha-glycoside of Neu5Ac. Thus, differentiation between the types of the SA-Gal linkage by the A viruses appeared to depend, at least partially, upon the recognition by the HA of the first hydrocarbon group of the aglycon. Type B virus strains were distinct in having a lower affinity for the Neu5Ac moiety and in providing a higher contribution of the asialic portions of sialosides to the HA-ligand interactions. The last effects are presumably due to the amino acid insertions in the type B HA surrounding the RBS, which makes the receptor-binding pocket deeper. The results obtained in the present investigation indicate that while the functional groups of Neu5Ac studied are recognized by the RBSs of all influenza viruses, the magnitude of their contribution to the binding energy, as well as the contribution of the asialic portion of the receptor, may vary in dependence upon the virus type, subtype, and strain.
Assuntos
Hemaglutininas Virais/metabolismo , Receptores de Superfície Celular , Receptores Imunológicos/metabolismo , Receptores Virais/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Ligação Competitiva , Sequência de Carboidratos , Ácidos Carboxílicos/metabolismo , Hemaglutininas Virais/química , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Molecular , Ácido N-Acetilneuramínico , Oxigênio/metabolismo , Receptores Imunológicos/química , Receptores Virais/química , Homologia de Sequência de Aminoácidos , Ácidos Siálicos/metabolismo , Sialoglicoproteínas/metabolismoRESUMO
A combined 1H-NMR and molecular mechanics study of [Cpp1, Sar7]AVP was performed in order to select the most probable conformations in DMSO solutions. The NMR constraints obtained were employed in the selection of starting conformations of the cyclic moiety of the analog. In particular, the diminished accessibility of the Asn5 NH proton to solvent and the close contact between Cpp1 and Cys6 C alpha H protons suggests a beta-turn conformation at the Phe3-Gln4 residues. Energy minimization was carried out both in the ECEPP/2 (rigid-valence geometry) and in the AMBER (flexible-valence geometry) force fields. Comparison of the experimental and calculated values of NMR characteristics has revealed that conformations containing type I, II, and III beta-turns at the Phe3-Gln4 residues are in reasonable agreement with the experimental data, with a dynamic equilibrium between the beta I (beta III) and beta II type structures of the cyclic part being the most probable. All of these conformations prefer the negative chirality of the disulfide bridge (theta 3 approximately -90 degrees). Five representative conformations were chosen for the acyclic tail: one with a beta I, one with a beta II'-turn at the Sar7-Arg8 residues, two extended-type conformations, and a conformation with a gamma-turn at Sar7. Because only high-energy extended conformations were in agreement with NMR data, it was concluded that the acyclic tail has considerable conformational flexibility in solution. The conformations obtained are discussed in terms of the structure-function relationship of the neurohypophyseal hormone analogs.
Assuntos
Arginina Vasopressina/análogos & derivados , Sequência de Aminoácidos , Arginina Vasopressina/química , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Molecular , Conformação Proteica , TermodinâmicaRESUMO
Sets of low-energy backbone conformations of the active tetragastrin analogue Boc-Trp-Leu-Asp-Phe-NH2 and two competitive antagonists Boc-Trp-Leu psi (CH2NH)-Asp-Phe-NH2 and Boc-Trp-Leu-Asp-O-CH2-CH2-C6H5 were obtained using theoretical conformational analysis methods. Groups of the conformations were selected for the three analogues, allowing a spatial matching of Trp, Asp and Phe residues responsible for the gastrin receptor binding. Three conformations possessing the lowest energies among the geometrically similar structures of these three peptides are suggested as a model for the "receptor-bound" conformations of these analogues. Backbone spatial folding resembling an alpha-helix turn is characteristic of these conformations. The correspondence of the proposed model to the available data on structure--activity relationships for tetragastrin analogues is discussed. Orientations of the putative receptor-bound conformations in a "water--lypophylic medium" two-phase system were investigated.
Assuntos
Tetragastrina/análogos & derivados , Sequência de Aminoácidos , Dados de Sequência Molecular , Proglumida/metabolismo , Conformação Proteica , Receptores da Colecistocinina/metabolismo , Relação Estrutura-Atividade , Tetragastrina/metabolismoRESUMO
Theoretical conformational analysis was carried out for several tetrapeptide analogues of beta-casomorphin and dermorphin containing a Phe residue in position 3. Sets of low-energy backbone structures of the mu-selective peptides [N-Me-Phe3, D-Pro4]-morphiceptin and Tyr-D-Orn-Phe-Asp-NH2 were obtained. These sets of structures were compared for geometrical similarity between themselves and with the low-energy conformations found for the delta-selective peptide Tyr-D-Cys-Phe-D-Pen-OH and nonactive peptide Tyr-Orn-Phe-Asp-NH2. Two pairs of geometrically similar conformations of mu-selective peptides, sharing no similarity with the conformations of peptides showing low affinity to the mu-receptor, were selected as two alternative models of probable mu-receptor-bound backbone conformations. Both models share geometrical similarity with the low-energy structures of the linear mu-selective peptide Tyr-D-Ala-Phe-Phe-NH2. Putative binding conformations of Tyr1 and Phe3 side chains are also discussed.
Assuntos
Encefalinas/química , Oligopeptídeos/química , Fenilalanina , Receptores Opioides/metabolismo , Sequência de Aminoácidos , Endorfinas/química , Endorfinas/metabolismo , Dados de Sequência Molecular , Estrutura Molecular , Oligopeptídeos/metabolismo , Peptídeos Opioides , Conformação Proteica , Receptores Opioides mu , Relação Estrutura-AtividadeRESUMO
Models of mu- and delta-receptor-bound backbone conformations of enkephalin cyclic analogues containing Phe4 were determined by comparing geometrical similarity among the previously found low-energy backbone structures of [D-Cys2,Cys5]-enkephalinamide, [D-Cys2,D-Cys5]-enkephalinamide, [D-Pen2,L-Pen5]-enkephalin and [D-Pen2,D-Pen5]-enkephalin. The present mu-receptor-bound conformation resembles a beta-I bend in the peptide backbone centred on the Gly3-Phe4 region. Two slightly different models were found for the delta-receptor-bound conformation; both of them are more extended than the mu-receptor-bound conformation and include a gamma-turn (or a gamma-like turn) on the Gly3 residue. Energetically favourable rotamers of Tyr and Phe side chains were also determined for the mu- and delta-conformations. The present models of mu- and delta-conformations share geometrical similarity with the low-energy structures of Leu-enkephalin and the Tyr-D-Lys-Gly-Phe-analogue.
Assuntos
Encefalinas/metabolismo , Receptores Opioides/metabolismo , Sequência de Aminoácidos , Encefalinas/química , Modelos Moleculares , Dados de Sequência Molecular , Conformação Proteica , Receptores Opioides delta , Receptores Opioides mu , Relação Estrutura-Atividade , TermodinâmicaRESUMO
Low-energy peptide backbone conformers were found by means of energy calculation for several cyclic analogues of enkephalin in an attempt to assess models for receptor-bound conformations for opioid receptors of the mu- and delta-types. They included [D-Cys2, L-Cys5]- and [D-Cys2, D-Cys5]-enkephalinamides showing moderate preference for mu-receptors, the delta-selective compounds [D-Pen2, L-Pen5] and [D-Pen2, D-Pen5]-enkephalins and Tyr-D-Lys-Gly-Phe- analogue possessing very high affinity to receptors of the mu-type. The low-energy conformers obtained for these analogues were in good agreement with the results of calculations by other authors and with experimental evidence. All of the analogues contain a Phe residue in position 4 of the peptide chain which facilitates the eventual search for geometrical similarity between the low-energy backbone conformers of different analogues in question.
Assuntos
Encefalinas/metabolismo , Fenilalanina , Receptores Opioides/metabolismo , Sequência de Aminoácidos , Animais , Dados de Sequência Molecular , Estrutura Molecular , Conformação Proteica , Ratos , TermodinâmicaRESUMO
The spatial structure of [des-Arg9]bradykinin, a selective agonist of the B1 type kinin receptors, was studied by means of theoretical conformational analysis. In order to restrict the number of conformations under consideration, we used available data on 1H NMR in (CD3)2SO solution indicating the C-terminal carboxyl group to interact with the guanidine group of Arg1 and the hydroxyl group of Ser6. The calculated set of [des-Arg9]bradykinin low-energy conformations was compared with the 1H NMR data. Four types of conformations were selected, which are consistent with experimental data and serve as sterically reliable models for the preferable three-dimensional structure of [des-Arg9]bradykinin in solution.
Assuntos
Bradicinina/análogos & derivados , Sequência de Aminoácidos , Dados de Sequência Molecular , Conformação Proteica , Soluções , Relação Estrutura-AtividadeRESUMO
Low-energy peptide backbone structures of dermorphin (DM), amide of its N-terminal pentapeptide (DM 1-5) and DM 1-5 analogues with substitutions of Gly4 for Leu, D-Gln, Aal or Tal were determined by energy calculations. The above analogues were shown to possess different affinities toward opiate receptors of mu-type. The comparison of low-energy backbone structures of DM, DM 1-5 and its analogues resulted in development of the dermorphin "biologically active" conformation being characteristic of its binding with mu-type receptors. The specific binding of dermorphin to this receptor apparently depends on the conformation of the whole N-terminal pentapeptide.
Assuntos
Oligopeptídeos/metabolismo , Sequência de Aminoácidos , Dados de Sequência Molecular , Peptídeos Opioides , Conformação Proteica , Receptores Opioides/metabolismo , Receptores Opioides mu , Relação Estrutura-AtividadeRESUMO
The three-dimensional structure of the cyclic analogs of bradykinin and substance P C-terminal hexapeptide was studied using conformational energy calculations. Initial conformations for energy minimization were selected with the aid of the measured intensities of local nuclear Overhauser effects (NOEs) and other 1H-NMR data. Expected values of the 1H-NMR parameters for low-energy conformations of the cyclopeptides were calculated and compared with those observed experimentally using semiquantitative gradation of NOE intensities. Several low-energy structures of the cyclic bradykinin analog, possessing similar backbone conformations stabilized by two beta-turns, are in agreement with experimental data. None of the low-energy conformations of the substance P cyclic hexapeptide were in satisfactory agreement with the experimental set of NOEs. The agreement was achieved only by averaging of the calculated 1H-NMR parameters over several combinations of the low-energy conformations.
Assuntos
Bradicinina/análogos & derivados , Peptídeos Cíclicos , Conformação Proteica , Substância P/análogos & derivados , Espectroscopia de Ressonância Magnética/métodos , Modelos Moleculares , TermodinâmicaRESUMO
2D 1H-NMR spectra of des-Gly9-[Arg8]vasopressin in dimethylsulfoxide have been taken and the 1H resonances have been assigned. The coupling constants and amide proton temperature coefficients (delta delta/delta T) have been measured and the NOE cross-peaks in the NOESY spectrum have been analyzed. The most essential information on the spatial structure of des-Gly9-[Arg8]vasopressin is extracted from the low delta delta/delta T value for Asn5 amide proton and from the NOE between the Cys1 and Cys6 alpha-protons. A diminished accessibility of the Asn5 NH proton for the solvent is ascribed to the presence of a beta-turn in the fragment 2-5. The distance between the Cys1 and Cys6 C alpha H protons seems to be less than 4 A. These constraints were taken into account in the conformational analysis of the title peptide. The derived set of the low-energy backbone conformations was analyzed against the background of the all available NMR data. The most probable conformation of the cyclic moiety in des-Gly9-[Arg8]vasopressin was found to be the type III beta-turn. The corner positions are occupied by the residues 3, 4, while the residues 1-2 and 5-6 are at the extended sites. Some NMR data indicate that this structure is in a dynamic equilibrium with other minor conformers.
Assuntos
Arginina Vasopressina/análogos & derivados , Sequência de Aminoácidos , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Conformação ProteicaRESUMO
H NMR resonances of [cyclo (9----18) Lys1, Gly6]bradykinin (CBK) in (CD3)2SO and H2O solution have been assigned by combined analysis of two-dimensional COSY and NOESY spectra. The presence of two slowly interchangeable conformers of CBK in (CD3)2SO is established, the minor conformer not exceeding 15% in the population. The minor conformer is absent from the aqueous solution, chemical shifts of the CBK and bradykinin NH and C alpha H protons differ insignificantly. The major CBK conformer contains at least two X-Pro trans-peptide groups and three amide protons NH Phe5, NH Arg9 and N zeta H Lys1 protected from solvent. A system of cross-peaks from the NOESY spectra of CBK in (CD3)2SO has been analysed and the maximum distance between backbone protons and neighbouring amino acid residues evaluated. The experimental data agree well with the assumed type II beta-bend in the sequence Pro2-Pro3-Gly4-Phe5. Spatial structure models for the backbone fragment 6-9 of CBK containing two intramolecular hydrogen bonds that involve the NH Arg9 and N zeta H Lys1 protons and the carbonyl groups of Phe5 and Gly4 are proposed.