Melanins From Opioid Peptides

Opioid peptides and other Tyr-NH₂-terminal peptides are substrates in vitro for mushroom and sepia tyrosinase, giving rise to synthetic melanins retaining the peptide moiety (opiomelanins). The melanopeptides are characterized by a total solubility in hydrophylic solvents at neutral and basic pH. Opioid peptides (enkephalins, endorphins, and esorphins), if oxidized by tyrosinase in the presence of Dopa, are easily incorporated into Dopa-melanin, producing mixed-type pigments that can also be solubilized in hydrophylic solvents. Melanins derived from opioid peptides exhibit paramagnetism, as evidenced by an EPR spectrum identical to that of Dopa-melanin. However, the presence of the linked peptide chain is able to influence dramatically the electron transfer properties and the oxidizing behaviour of the melanopeptides, so that whereas Tyr-Gly-melanin appears to behave as Dopa-melanin, Enk-melanin does not exhibit any oxidizing activity. Opiomelanins are characterized by a peculiar UV-VIS spectrum; that is, by the presence of a distinct peak (330 nm) that disappears upon chemical treatment by acid hydrolysis. Opiomelanins are stable pigments at neutral and basic pH in the dark, whereas the addition of H₂O₂ leads to a 15% degradation. Under simulated solar illumination, opiomelanins are more easily destroyed with respect to Dopa-melanin, with increasing degradation when exposed to increased hydrogen peroxide concentrations and more alkaline pH. Some speculations on the possible existence and role of opiomelanins have been outlined.     

Regulation of Opioid Receptors by Their Endogenous Opioid Peptides

Cellular and Molecular Neurobiology - Activation of μ, δ, and κ opioid receptors by endogenous opioid peptides leads to the regulation of many emotional and physiological responses....     

Peptides from Fish By-product Protein Hydrolysates and Its Functional Properties: an Overview

The inadequate management of fish processing waste or by-products is one of the major problems that fish industry has to face nowadays. The mismanagement of this raw material leads to economic loss and environmental problems. The demand for the use of these by-products has led to the development of several processes in order to recover biomolecules from fish by-products. An efficient way to add value to fish waste protein is protein hydrolysis. Protein hydrolysates improve the functional properties and allow the release of peptides of different sizes with several bioactivities such as antioxidant, antimicrobial, antihypertensive, anti-inflammatory, or antihyperglycemic among others. This paper reviews different methods for the production of protein hydrolysates as well as current research about several fish by-products protein hydrolysates bioactive properties, aiming the dual objective: adding value to these underutilized by-products and minimizing their negative impact on the environment.     

细胞穿膜肽入胞检测方法概述

迄今为止,已有多达上百种的细胞穿膜肽(cell-penetrating peptides,CPPs)被发现报道,但这类多肽分子的入胞能力参差不齐,限制了其作为药物载体的应用。虽然已有多种实验方法可用于细胞穿膜肽入胞的检测,但由于缺乏通用的技术来确切证实CPPs的入胞能力,所以应当结合使用多种方法以降低误差。对不同的技术在检测CPPs入胞时的优缺点进行比较,并针对性地提出比较理想的解决方案,可为制订CPPs入胞标准化检测步骤提供一些参考。     

Consequences of Cis-amide Bond Simulation in Opioid Peptides

Six analogs of leucine-enkephalin were synthesized in which a 1,5-disubstituted tetrazole ring was incorporated in order to lock selected peptide bonds in cis geometry. The obtained compounds were examined based on their biological effects in vivo and in vitro. Only one analog was completely inactive in binding assays being very weakly active in the antinociceptive test. The remaining five compounds displayed at least weak receptor affinity and in vivo activity.     

Role of Opioid Peptides in Behavior of Invertebrates

Opioid peptides have been revealed in representatives of practically all large taxonomic groups of invertebrates, and the opiate receptors are found even in unicellulars. The opioid system seems to belong to the evolutionary ancient signal systems. The comparative data indicate that the most conservative and ancient function of opioids is control of the adequate level of protective reactions. In the infusorian Stentor the opiate ligands suppress a contractile response to mechanical stimulation, i.e., the protective behavior. In all studies multicellular invertebrates, agonists also suppress protective behavior, whereas antagonists produce opposite effects. This initially signal meaning of opioids might have become a basis for divergent development of their functions in evolution. Already in higher invertebrates, molluscs and arthropods, many functions of opioids, for example, stress-induced analgesia, regulation of feeding and mating behavior, of social aggression, are similar to those in vertebrates. It is suggested that the main events in formation of functions of the endogenous opioid system have occurred in the lower invertebrates that have remained so far the least studied.     

Opioid Peptides: Synthesis and Biological Activity of New Endomorphin Analogues

Summary Syntheses are described of new endomorphin 1 and 2 peptoid–peptide hybrids in which Tyr¹ and either one or both Phe³ and Phe⁴ have been replaced by N-substituted-glycine. The preparation is also described of two glycosylated Hyp²-endomorphin 2 analogues in which either 2,3,4,6-tetra-O-acetyl glucose or glucose are β-O-glycosidically linked to the hydroxyproline residue. The Hyp²-endomorphin sequences have also been elongate by adding a C-terminal β-alanine residue and several linear dimers have been prepared by coupling either the native peptides or the modified analogues. The cyclo endomorphin 2 has also been synthesized. Preliminary pharmacological experiments on isolated organ preparations showed that the agonist activities of both endomorphin 1 and 2 are not significantly affected by the Pro/Hyp substitution. Phe⁴/Nphe substitution in the endomorphin 1 reduced the potency on guinea pig ileum (GPI) by about 100 times and abolished the agonist activity on mouse vas deferens (MVD) preparation. The decrease of the agonist activity induced by modification of one phenylalanine residue only, either Phe³ or Phe⁴, is lower on endomorphin 2. Either modification of both Phe³ and Phe⁴ or glycosylation of the Hyp²-endomorphin 2 cancelled any agonist activity on both preparations. The linear peptide dimers [endomorphin 1]₂, [endomorphin 2]₂, [Hyp²-endomorphin 1]₂, [Hyp²-endomorphin 2]₂, [Hyp²-endomorphin 1-Hyp²-endomorphin 2]₂ or [Hyp²-endomorphin 2-Hyp²-endomorphin 1]₂, are 7–19 times less potent than endomorphin 1 on GPI and significantly less active than endomorphins 1 and 2 on MVD. The other afforded modifications significantly affected or abolished the agonist activity of the resulting endomorphin analogues on both GPI and MVD preparations.The α-amino acid residues are of the L-configuration. Standard abbreviations for amino acid derivatives and peptides are according to the suggestions of the IUPAC-IUB Commission on Biochemical Nomenclature (1984) Eur. J. Biochem., 138, 9–37. Abbreviations listed in the guide published in (2003) J. Peptide Sci., 9, 1–8 are used without explanation.     

Consequences of Cis-amide Bond Simulation in Opioid Peptides

Summary Six analogs of leucine-enkephalin were synthesized in which a 1,5-disubstituted tetrazole ring was incorporated in order to lock selected peptide bonds in cis geometry. The obtained compounds were examined based on their biological effects in vivo and in vitro. Only one analog was completely inactive in binding assays being very weakly active in the antinociceptive test. The remaining five compounds displayed at least weak receptor affinity and in vivo activity.     

Functional Roles of Gangliosides in Neurodevelopment: An Overview of Recent Advances

Gangliosides are sialic acid-containing glycosphingolipids that are most abundant in the nervous system. They are localized primarily in the outer leaflets of plasma membranes and participated in cell–cell recognition, adhesion, and signal transduction and are integral components of cell surface microdomains or lipid rafts along with proteins, sphingomyelin and cholesterol. Ganglioside-rich lipid rafts play an important role in signaling events affecting neural development and the pathogenesis of certain diseases. Disruption of gangloside synthase genes in mice induces developmental defects and neural degeneration. Targeting ganglioside metabolism may represent a novel therapeutic strategy for intervention in certain diseases. In this review, we focus on recent advances on metabolic and functional studies of gangliosides in normal brain development and in certain neurological disorders.     

The Effects of Opioid Peptides on Dopamine Release in the Nucleus Accumbens: An In Vivo Microdialysis Study

An involvement of the mesolimbic dopamine (DA) system in mediating the motivational effects of opioids has been suggested. Accordingly, the present study employed the technique of in vivo microdialysis to examine the effects of selective mu-, delta-, and kappa- opioids on DA release in the nucleus accumbens (NAC) of anesthetized rats. Microdialysis probes were inserted into the NAC and perfusates were analyzed for DA and its metabolites, dihydroxyphenylacetic acid (DO-PAC) and homovanillic acid (HVA), using a reverse-phase HPLC system with electrochemical detection for separation and quantification. Intracerebroventricular (i.c.v.) administration of selective mu-opioid [D-Ala2, N-methyl-Phe4, Gly5-ol]-enkephalin (DAMGO) or delta-opioid [D-Pen2, D-Pen5]-enkephalin (DPDPE) agonists, at doses that function as positive reinforcers in rats, resulted in an immediate and significant increase in extracellular DA. DOPAC and HVA levels were also significantly increased. The effects of DAMGO were blocked by the selective mu-antagonist D-Pen-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2 (CTOP) whereas those of DPDPE were blocked by the delta-antagonist allyl2-Tyr-Aib-Aib-Phe-Leu-OH (ICI 174,864). In contrast to mu- and delta-agonists, the kappa-agonist N-CH3-Tyr-Gly-Gly-Phe-Leu-Arg-N-CH3-Arg-D-Leu-NHC2H5 (E-2078), a dynorphin analog that produces aversive states, decreased DA release in a biphasic manner. Norbinaltorphimine, a selective kappa-antagonist, could block this effect. These results demonstrate that mu-, delta-, and kappa-opioid agonists differentially affect DA release in the NAC and this action is centrally mediated.     

Isolation and Characterization of Opioid Peptides from Rabbit Cerebellum

The rabbit cerebellum has been shown to contain significant quantities of opioid receptors consisting of both mu- and kappa-subtypes. To determine the nature of the endogenous opioid ligands in this tissue, extracts from rabbit cerebellum were separated by various chromatography techniques and fractions were assayed initially for opioid peptides with a radioimmunoassay capable of detecting all peptides with an amino-terminal Tyr-Gly-Gly-Phe sequence. This sequence is common to all mammalian opioid peptides and is critical for recognition by all known opioid receptors. Each of the three immunoreactive opioid peptide peaks detected was purified to homogeneity and subjected to amino acid composition and sequence analysis. One peak was analyzed further by mass spectrometry. This identified the major opioid peptides in the cerebellum as [Met5]enkephalin, [Leu5]enkephalin, and heptapeptide [Met5]enkephalyl-Arg6-Phe7. The comprehensiveness of this initial detection scheme in identifying biologically active opioid peptides was substantiated through subsequent analysis. Using specific radioimmunoassays for representative opioid peptides of the three opioid systems currently known, no other peptides of either the proenkephalin, proopiomelanocortin, or prodynorphin series were detected in any appreciable amounts. Collectively, these results are consistent with the position that rabbit cerebellar opioids are derived from proenkephalin. However, given that no appreciable quantities of either [Met5]enkephalyl-Arg6-Arg7-Val8-NH2 (metorphamide) or [Met5]enkephalyl-Arg6-Gly7-Leu8 were detected suggests that rabbit proenkephalin may have a slightly altered sequence and/or is differentially processed relative to other mammalian species studied.     

半导体激光针对中枢及外周内阿片肽神经递质的影响

本工作应用半导体激光６５０ｎｍ １０ｍＷ ＣＷ照射“扶突”穴，发现Ｌ－ＥＮＫ除在尾核区有非常显著的增加外，而在海马，丘脑，下丘脑则都有不同程度的下降，其显著性Ｐ值分别为Ｐ〈０．０５，Ｐ〈０；０１，Ｐ〈０．０１。在外周血浆中Ｌ－ＥＮＫ则亦有显著的增加。     

The Relationship Between Structure and Activity Among Opioid Peptides

Since the discovery and isolation of the endogenous opioid peptides Leu- and Met-enkephalin, structural studies have been focused on deducing the bioactive conformation of the peptide ligands. Theoretically, linear peptides can have many different backbone conformations, yet early X-ray studies on enkephalin and its analogues showed only two different backbone conformations: extended and single -bend. More recent reports include a third conformation for Leu-enkephalin and constrained opioid peptides from two new classes (i.e. cyclic and all-aromatic peptides). In this report the relationship between solid-state X-ray structure and opioid peptide activity is examined. The N-terminal amine nitrogen and the two aromatic rings have previously been identified as structural features important to the biological activity of opioid peptides. From X-ray studies we find that the distances between the centroids of the aromatic rings, and between the N-terminal amine nitrogen and the centroid of the phenylalanine ring, vary over a large range. There is a discernible relationship, however, between the separation of the two rings and their orientation that correlates with activity.     

Reaction of Opioid Peptides with Neutral Endopeptidase (''Enkephalinase")

The kinetics of the reactions of nine opioid peptides with the neutral endopeptidase ("enkephalinase") activities of human kidney, rat kidney, and rat brain have been determined. These opioid peptides can be divided into two classes, those that are good inhibitors of Leu5-enkephalin hydrolysis (Ki less than 75 microM) and good substrates for the enzyme, and those that are poor inhibitors (Ki greater than 500 microM) and are not substrates for the enzyme. The former group includes Leu5-enkephalin, Met5-enkephalin, Met5-enkephalin-Arg6-Phe7, beta-lipotropin, and gamma-endorphin, while the nonreactive opioid peptides include alpha-neo-endorphin, beta-neo-endorphin, dynorphin, and beta-endorphin. These results suggest that those peptides containing the Met5-enkephalin sequence are more reactive than those containing the Leu5-enkephalin sequence. The lack of specificity of this neutral endopeptidase indicates that it may function in the degradation of a variety of biologically active peptides.     

The Relationship Between Structure and Activity Among Opioid Peptides

Summary Since the discovery and isolation of the endogenous opioid peptides Leu- and Met-enkephalin, structural studies have been focused on deducing the bioactive conformation of the peptide ligands. Theoretically, linear peptides can have many different backbone conformations, yet early, X-ray studies on enkephalin and its analogues showed only two different backbone conformations: extended and single β-bend. More recent reports include a third conformation for Leu-enkephalin and constrained opioid peptides from two ‘new’ classes (i.e. cyclic and ‘allaromatic’ peptides). In this report the relationship between solid-state X-ray structure and opioid peptide activity is examined. The N-terminal amine nitrogen and the two aromatic rings have previously been identified as structural features important to the biological activity of opioid peptides. From X-ray studies we find that the distances between the centroids of the aromatic rings, and between the N-terminal amine nitrogen and the centroid of the phenylalanine ring, vary over a large range. There is a discernible relationship, however, between the separation of the two rings and their orientation that correlates with activity.     

Subcellular Distribution of Opioid Peptides in Rat Hypothalamus and Pituitary

Homogenates of rat anterior lobe (AL) and neurointermediate lobe (NIL) pituitary and rat hypothalamus were subjected to subcellular fractionation and density gradient centrifugation. The subcellular distribution of immunoreactive dynorophin A (ir-Dyn A) in NIL was found to be similar to that of ir-arginine vasopressin (ir-AVP). ir-Dyn A migrated as a discrete band on sucrose density gradients, which corresponded in sedimentation rate to that of ir-AVP, suggesting that these two peptides are stored within organelles of similar size and density. Two other products of prodynorphin, ir-alpha-neoendorphin (ir-alpha-nEND) and ir-Dyn A-(1-8) also comigrated with ir-AVP. ir-[Leu5]-enkephalin (ir-LE), which may be a product of prodynorphin or proenkephalin, was also found to migrate in this region of the gradient. When a homogenate of rat hypothalamus was prepared using a method that has been developed for synaptosome isolation, ir-Dyn A was found to comigrate with Na+/K+-activated adenosine triphosphatase (Na/K-ATPase), a synaptosomal marker enzyme. Using a more concentrated homogenate ir-Dyn A was found to migrate to a less dense region where peptide-containing synaptic vesicles have previously been localized. When a synaptosomal preparation was lysed in hypotonic solution a shift was seen in the migration rate of ir-Dyn A to this region of the gradient (containing putative synaptic vesicles). Thus the bulk of hypothalamic dynorphin appears to be present within synaptosome-like structures which, upon lysis, release a less dense, smaller subcellular organelle corresponding in sedimentation characteristics to other types of peptide-containing synaptic vesicles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Neuroblastoma × Glioma Hybrid Cells Synthesize Enkephalin-Like Opioid Peptides

Partially purified extracts from neuroblastoma X glioma hybrid cells 108CC15 inhibit, like opioids, the prostaglandin E1-evoked formation of cyclic AMP in a dose-dependent manner in the same hybrid cells. The inhibition is prevented by the opioid antagonist naloxone. In addition, the same extract competes with [3H]naloxone and [3H]Leu-enkephalin for binding to opioid receptors of hybrid cell membranes and to a specific antiserum, respectively. The opioid activity in the extracts is destroyed by carboxypeptidase A and leucine aminopeptidase, but not by trypsin. Further purification of the extracts by HPLC, TLC, or high-voltage paper electrophoresis reveals in each case two active fractions which behave like Met- and Leu-enkephalin. The Met-enkephalin-like, but not the Leu-enkephalin-like, fraction is inactivated by treatment with BrCN. Dimethylaminonaphtylsulfonyl (dansyl) derivatives of Met- and Leu-enkephalin correspond to [3H]dansyl derivatives of Met-like substances from hybrid cells. Three to four times as much Met-enkephalin-like as Leu-enkephalin-like material is present in the extract. The overall concentration of opioid peptides in the hybrid cells varies between 0.03 and 1.0 pmol Leu-enkephalin equivalents per mg protein. The amount of opioids in the hybrid cells is strongly dependent on the cell density. The findings suggest that neuroblastoma X glioma hybrid cells contain opioid peptides that are very similar, if not identical, to Met- and Leu-enkephalin. Opioid activity can also be detected in other neuronal cell lines and even in glioma cells.     

Electrically-Induced Release of Opioid Peptides from the Guinea-Pig Myenteric Plexus Preparation

Abstract

Preparations of guinea-pig myenteric plexus-longitudinal muscle suspended in Krebs solution were stimulated electrically in the presence of cycloheximide and tetraethylammonium. The amounts of eleven endogenous opioid peptides released into the perifusing Krebs solution were determined and correlated with the decrease in the tissue contents induced by stimulation. For pro-enkephalin fragments the ratio of release to reduction in tissue contents was 29 to 43% for [Met]enkephalin, [Leu]enkephalin, [Met]enkephalyl-RF and [Met]enkephalyl-RGL. With [Met]enkephalyl-RRV-NH₂ (BAM-8) the ratio was higher by 50% or more. However, it is of interest that there was no release of the probable precursor [Met]enkephalyl-RRVGRPEWWMDYQ(BAM-18). In this context it may be important that BAM-8 is the only endogenous opioid peptide having-NH₂ at the C-terminal. The low tissue levels of pro-dynorphin derived peptide have made estimation of release unreliable.     

Urban Folk Medicine: A Functional Overview

Social and economic functions of folk illness and folk medicine are fairly well known for rural contexts but still lack codification for urban milieus. In this exploratory paper, folk health practices are examined in terms of their response to urban socioeconomic characteristics. Such practices appear to serve functions of acculturation, guilt displacement resulting from failure to achieve, and subgroup identity maintenance, among others. Folk practices are resilient, readily shifting to adjunct functions of healing under pressure from effective modern medical and welfare systems . [medical anthropology, urban anthropology, curanderismo, ethnology, health]