RESUMEN
Somatostatin receptor 5 (SSTR5) is an important G protein-coupled receptor and drug target for neuroendocrine tumors and pituitary disorders. This study presents two high-resolution cryogenicelectron microscope structures of the SSTR5-Gi complexes bound to the cyclic neuropeptide agonists, cortistatin-17 (CST17) and octreotide, with resolutions of 2.7 Å and 2.9 Å, respectively. The structures reveal that binding of these peptides causes rearrangement of a "hydrophobic lock", consisting of residues from transmembrane helices TM3 and TM6. This rearrangement triggers outward movement of TM6, enabling Gαi protein engagement and receptor activation. In addition to hydrophobic interactions, CST17 forms conserved polar contacts similar to somatostatin-14 binding to SSTR2, while further structural and functional analysis shows that extracellular loops differently recognize CST17 and octreotide. These insights elucidate agonist selectivity and activation mechanisms of SSTR5, providing valuable guidance for structure-based drug development targeting this therapeutically relevant receptor.
Asunto(s)
Octreótido , Receptores de Somatostatina , Receptores de Somatostatina/metabolismo , Receptores de Somatostatina/agonistas , Receptores de Somatostatina/química , Humanos , Octreótido/química , Octreótido/farmacología , Octreótido/metabolismo , Neuropéptidos/metabolismo , Neuropéptidos/química , Microscopía por Crioelectrón , Unión Proteica , Péptidos Cíclicos/química , Péptidos Cíclicos/farmacología , Péptidos Cíclicos/metabolismo , Somatostatina/metabolismo , Somatostatina/química , Somatostatina/análogos & derivados , Modelos Moleculares , Células HEK293RESUMEN
Neuropeptides represent a unique class of signaling molecules that have garnered much attention but require special consideration when identifications are gleaned from mass spectra. With highly variable sequence lengths, neuropeptides must be analyzed in their endogenous state. Further, neuropeptides share great homology within families, differing by as little as a single amino acid residue, complicating even routine analyses and necessitating optimized computational strategies for confident and accurate identifications. We present EndoGenius, a database searching strategy designed specifically for elucidating neuropeptide identifications from mass spectra by leveraging optimized peptide-spectrum matching approaches, an expansive motif database, and a novel scoring algorithm to achieve broader representation of the neuropeptidome and minimize reidentification. This work describes an algorithm capable of reporting more neuropeptide identifications at 1% false-discovery rate than alternative software in five Callinectes sapidus neuronal tissue types.
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Algoritmos , Bases de Datos de Proteínas , Neuropéptidos , Programas Informáticos , Neuropéptidos/análisis , Neuropéptidos/química , Animales , Espectrometría de Masas/métodos , Secuencia de Aminoácidos , Proteómica/métodos , Espectrometría de Masas en Tándem/métodosRESUMEN
BACKGROUND: Larval settlement and metamorphosis represent critical events in the life history of marine benthic animals. Myoinhibitory peptide (MIP) plays a pivotal role in larval settlement of marine invertebrates. However, the molecular mechanisms of MIP involved in this process are not well understood. RESULTS: In this study, we evaluated the effects of thirteen MIP mature peptides on triggering the larval settlement of Urechis unicinctus (Xenopneusta, Urechidae), and determined that MIP2 was the principal neuropeptide. Transcriptomic analysis was employed to identify differentially expressed genes (DEGs) between the MIP2-treated larvae and normal early-segmentation larvae. Both cAMP and calcium signaling pathways were enriched in the DEGs of the MIP2-treated larvae, and two neuropeptide receptor genes (Spr, Fmrfar) were up-regulated in the MIP2-treated larvae. The activation of the SPR-cAMP pathway by MIP2 was experimentally validated in HEK293T cells. Furthermore, fourteen cilia-related genes, including Tctex1d2, Cfap45, Ift43, Ift74, Ift22, Cav1 and Mns1, etc. exhibited down-regulated expression in the MIP2-treated larvae. Whole-mount in situ hybridization identified two selected ciliary genes, Tctex1d2 and Cfap45, were specially expressed in circumoral ciliary cells of the early-segmentation larvae. Knocking down Tctex1d2 mRNA levels by in vivo RNA interference significantly increased the larval settlement rate. CONCLUSION: Our findings suggest that MIP2 inhibits the function of the cilia-related genes, such as Tctex1d2, through the SPR-cAMP-PKA pathway, thereby inducing larval settlement in U. unicinctus. The study contributes important data to the understanding of neuropeptide regulation in larval settlement.
Asunto(s)
Neuropéptidos , Poliquetos , Humanos , Animales , Larva/genética , Células HEK293 , Poliquetos/genética , Neuropéptidos/genética , Neuropéptidos/química , Perfilación de la Expresión GénicaRESUMEN
Neuritin plays an important role in promoting nerve injury repair and maintaining synaptic plasticity, making it a potential therapeutic target for the treatment of nerve injury and neurodegenerative diseases. The present study aimed to obtain an active, unlabeled neuritin protein. Initially, a neuritin protein expression system with an enterokinase site was constructed in Escherichia coli. After optimizing induction conditions and screening for high expression, a neuritin recombinant protein with purity exceeding 85 % was obtained through Ni-affinity chromatography. Subsequently, unlabeled neuritin with a molecular weight of 11 kDa was obtained through the enzymatic cleavage of the His label using an enterokinase. Furthermore, a neuritin recombinant protein with purity exceeding 95 % was obtained using gel chromatography. Functional investigations revealed that neurite outgrowth of PC12 cells was stimulated by the isolated neuritin. This study establishes a method to obtain active and unlabeled neuritin protein, providing a foundation for subsequent research on its biological functions.
Asunto(s)
Escherichia coli , Proteínas Ligadas a GPI , Proteínas Recombinantes , Animales , Células PC12 , Ratas , Proteínas Ligadas a GPI/genética , Proteínas Ligadas a GPI/química , Proteínas Ligadas a GPI/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/aislamiento & purificación , Escherichia coli/genética , Escherichia coli/metabolismo , Neuropéptidos/genética , Neuropéptidos/química , Neuropéptidos/metabolismo , Enteropeptidasa/metabolismo , Enteropeptidasa/genética , Enteropeptidasa/química , Cromatografía de Afinidad , Proyección Neuronal/efectos de los fármacos , Cromatografía en Gel , Expresión GénicaRESUMEN
CAPA neuropeptides regulate the diuresis/ antidiuresis process in insects by activating specific cognate receptor, CAPAr. In this study, we characterized the CAPAr gene (BtabCAPAr) in the whitefly, Bemisia tabaci Asia II 1. The two alternatively spliced isoforms of BtabCAPAr gene, BtabCAPAr-1 and BtabCAPAr-2, having six and five exons, respectively, were identified. The BtabCAPAr gene expression was highest in adult whitefly as compared to gene expression in egg, nymphal and pupal stages. Among the three putative CAPA peptides, CAPA-PVK1 and CAPA-PVK2 strongly activated the BtabCAPAr-1 with very low EC50 values of 0.067 nM and 0.053 nM, respectively, in heterologous calcium mobilization assays. None of the peptide activated the alternatively spliced isoform BtabCAPAr-2 that has lost the transmembrane segments 3 and 4. Significant levels of mortality were observed when whiteflies were fed with CAPA-PVK1 at 1.0 µM (50.0%), CAPA-PVK2 at 100.0 nM (43.8%) and CAPA-tryptoPK 1.0 µM (40.0%) at the 96 h after the treatment. This study provides valuable information to design biostable peptides to develop a class of insecticides.
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Hemípteros , Neuropéptidos , Animales , Péptidos/metabolismo , Neuropéptidos/química , Neuropéptidos/genética , Neuropéptidos/metabolismo , Transducción de Señal , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Hemípteros/genética , Hemípteros/metabolismoRESUMEN
In the year 2002, DNA loss model (DNA-LM) postulated that neuropeptide genes to emerged through codons loss via the repair of damaged DNA from ancestral gene namely Neuropeptide Precursor Predictive (NPP), which organization correspond two or more neuropeptides precursors evolutive related. The DNA-LM was elaborated according to amino acids homology among LWamide, APGWamide, red pigment-concentrating hormone (RPCH), adipokinetic hormones (AKHs) and in silico APGW/RPCH NPPAPGW/AKH NPP were proposed. With the above principle, it was proposed the evolution of corazonin (CRZ), gonadotropin-releasing hormone (GnRH), AKH, and AKH/CRZ (ACP), but any NPP never was considered. However, the evolutive relation via DNA-LM among these neuropeptides precursors not has been established yet. Therefore, the transcriptomes from crabs Callinectes toxotes and Callinectes arcuatus were used to characterized ACP and partial CRZ precursors, respectively. BLAST alignment with APGW/RPCH NPP and APGW/AKH NPP allow identified similar NPP in the rotifer Brachionus plicatilis and other invertebrates. Moreover, three bioinformatics algorithms and manual verification were used to purify 13,778 sequences, generating a database with 719 neuropeptide precursors. Phylogenetic trees with the DNA-LM parameters showed that some ACP, CRZ, AKH2 and two NPP share nodes with GnRH from vertebrates and some of this neuropeptide had nodes in invertebrates. Whereas the phylogenetic tree with standard parameters do not showed previous node pattern. Robinson-Foulds metric corroborates the differences among phylogenetic trees. Homology relationship showed four putative orthogroups; AKH4, CRZ, and protostomes GnRH had individual group. This is the first demonstration of NPP in species and would explain the evolution neuropeptide families by the DNA-LM.
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Hormona Liberadora de Gonadotropina , Neuropéptidos , Humanos , Animales , Hormona Liberadora de Gonadotropina/genética , Hormona Liberadora de Gonadotropina/metabolismo , Filogenia , Evolución Molecular , Neuropéptidos/genética , Neuropéptidos/química , Neuropéptidos/metabolismo , Invertebrados/genética , ADN/metabolismoRESUMEN
The field of peptidomics has been under development since its start more than 20 years ago. In this chapter we provide a personal outlook for future directions in this field. The applications of peptidomics technologies are spreading more and more from classical research of peptide hormones and neuropeptides towards commercial applications in plant and food-science. Many clinical applications have been developed to analyze the complexity of biofluids, which are being addressed with new instrumentation, automization, and data processing. Additionally, the newly developed field of immunopeptidomics is showing promise for cancer therapies. In conclusion, peptidomics will continue delivering important information in classical fields like neuropeptides and peptide hormones, benefiting from improvements in state-of-the-art technologies. Moreover, new directions of research such as immunopeptidomics will further complement classical omics technologies and may become routine clinical procedures. Taken together, discoveries of new substances, networks, and applications of peptides can be expected in different disciplines.
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Neuropéptidos , Hormonas Peptídicas , Proteómica/métodos , Neuropéptidos/químicaRESUMEN
CPN-116 is a peptidic agonist that activates human neuromedin U receptor type 2 (NMUR2) but suffers from chemical instability due to inherent backbone isomerization on the Dap residue. To address this, a Leu-Dap-type (Z)-chloroalkene dipeptide isostere was synthesized diastereoselectively as a surrogate of the Leu-Dap peptide bond to develop a (Z)-chloroalkene analogue of CPN-116. The synthesized CPN-116 analogue is stable in 1.0 M phosphate buffer (pH 7.4) without backbone isomerization and can activate NMUR2 with similar potency to CPN-116 at nM concentrations (EC50 = 1.0 nM).
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Neuropéptidos , Humanos , Neuropéptidos/química , Amidas/farmacología , Péptidos , Receptores de Neurotransmisores/agonistasRESUMEN
Neuropeptides not only work through nervous system but some of them also work peripherally to regulate numerous physiological processes. They are important in regulation of numerous physiological processes including growth, reproduction, social behavior, inflammation, fluid homeostasis, cardiovascular function, and energy homeostasis. The various roles of neuropeptides make them promising candidates for prospective therapeutics of different diseases. Currently, NeuroPep has been updated to version 2.0, it now holds 11,417 unique neuropeptide entries, which is nearly double of the first version of NeuroPep. When available, we collected information about the receptor for each neuropeptide entry and predicted the 3D structures of those neuropeptides without known experimental structure using AlphaFold2 or APPTEST according to the peptide sequence length. In addition, DeepNeuropePred and NeuroPred-PLM, two neuropeptide prediction tools developed by us recently, were also integrated into NeuroPep 2.0 to help to facilitate the identification of new neuropeptides. NeuroPep 2.0 is freely accessible at https://isyslab.info/NeuroPepV2/.
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Bases de Datos de Proteínas , Anotación de Secuencia Molecular , Neuropéptidos , Secuencia de Aminoácidos , Neuropéptidos/química , Anotación de Secuencia Molecular/métodosRESUMEN
The advent of the fourth industrial revolution, characterized by artificial intelligence (AI) as its central component, has resulted in the mechanization of numerous previously labor-intensive activities. The use of in silico tools has become prevalent in the design of biopharmaceuticals. Upon conducting a comprehensive analysis of the genomes of many organisms, it has been discovered that their tissues can generate specific peptides that confer protection against certain diseases. This study aims to identify a selected group of neuropeptides (NPs) possessing favorable characteristics that render them ideal for production as neurological biopharmaceuticals. Until now, the construction of NP classifiers has been the primary focus, neglecting to optimize these characteristics. Therefore, in this study, the task of creating ideal NPs has been formulated as a multi-objective optimization problem. The proposed framework, NPpred, comprises two distinct components: NSGA-NeuroPred and BERT-NeuroPred. The former employs the NSGA-II algorithm to explore and change a population of NPs, while the latter is an interpretable deep learning-based model. The utilization of explainable AI and motifs has led to the proposal of two novel operators, namely p-crossover and p-mutation. An online application has been deployed at https://neuropred.anvil.app for designing an ideal collection of synthesizable NPs from protein sequences.
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Algoritmos , Inteligencia Artificial , Humanos , Neuropéptidos/genética , Neuropéptidos/química , Diseño de Fármacos , Simulación por Computador , Aprendizaje ProfundoRESUMEN
Highly specialized cells, such as neurons and podocytes, have arborized morphologies that serve their specific functions. Actin cytoskeleton and its associated proteins are responsible for the distinctive shapes of cells. The mechanism of their cytoskeleton regulation - contributing to cell shape maintenance - is yet to be fully clarified. Inverted formin 2 (INF2), one of the modulators of the cytoskeleton, is an atypical formin that can both polymerize and depolymerize actin filaments depending on its molar ratio to actin. Prior work has established that INF2 binds to the sides of actin filaments and severs them. Drebrin is another actin-binding protein that also binds filaments laterally and stabilizes them, but the interplay between drebrin and INF2 on actin filament stabilization is not well understood. Here, we have used biochemical assays, electron microscopy, and total internal reflection fluorescence microscopy imaging to show that drebrin protects actin filaments from severing by INF2 without inhibiting its polymerization activity. Notably, truncated drebrin - DrbA1-300 - is sufficient for this protection, though not as effective as the full-length protein. INF2 and drebrin are abundantly expressed in highly specialized cells and are crucial for the temporal regulation of their actin cytoskeleton, consistent with their involvement in peripheral neuropathy.
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Actinas , Forminas , Neuropéptidos , Citoesqueleto de Actina/química , Actinas/química , Forminas/química , Neuropéptidos/química , Extensiones de la Superficie Celular/química , Neuronas/metabolismo , Microscopía ElectrónicaRESUMEN
Neuropeptides are involved in many biological processes in insects. However, it is unclear what role neuropeptides play in Spodoptera litura adaptation to phytochemical flavone. In this study, 63 neuropeptide precursors from 48 gene families were identified in S. litura, including two neuropeptide F genes (NPFs). NPFs played a positive role in feeding regulation in S. litura because knockdown of NPFs decreased larval diet intake. S. litura larvae reduced flavone intake by downregulating NPFs. Conversely, the flavone intake was increased if the larvae were treated with NPF mature peptides. The NPF receptor (NPFR) was susceptible to the fluctuation of NPFs. NPFR mediated NPF signaling by interacting with NPFs to regulate the larval diet intake. In conclusion, this study suggested that NPF signaling regulated diet intake to promote S. litura adaptation to flavone, which contributed to understanding insect adaptation mechanisms to host plants and provide more potential pesticidal targets for pest control.
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Proteínas de Insectos , Larva , Neuropéptidos , Spodoptera , Animales , Spodoptera/fisiología , Spodoptera/metabolismo , Neuropéptidos/metabolismo , Neuropéptidos/genética , Neuropéptidos/química , Larva/crecimiento & desarrollo , Larva/metabolismo , Larva/química , Proteínas de Insectos/metabolismo , Proteínas de Insectos/genética , Proteínas de Insectos/química , Flavonas/metabolismo , Flavonas/química , Conducta Alimentaria , Secuencia de AminoácidosRESUMEN
BACKGROUND: Neuropeptide pedal peptide (PP) and orcokinin (OK), which are structurally related active peptides, have been widely discovered in invertebrates and constitute the PP/OK neuropeptide family. They have complex structures and play myriad roles in physiological processes. To date, there have been no related reports of PP/OK-type neuropeptide in cephalopods, which possess a highly differentiated multi-lobular brain. METHODS: Rapid Amplification of cDNA Ends (RACE) was employed to obtain the open reading frame (ORF) of PP/OK-type neuropeptide in Sepiella japonica (termed as Sj-PP/OK). Various software were used for sequence analysis. Semi-quantitative PCR was applied to analyze the tissue distribution profile, quantitative real-time PCR (qRT-PCR) was used to study spatio-temporal expression throughout the entire growth and development period, and in situ hybridization (ISH) was employed to observe the tissue location of Sj-PP/OK. RESULTS: in the present study, we identified the ORF of Sj-PP/OK. The putative precursor of Sj-PP/ OK encodes 22 mature peptides, of which only tridecapeptides could undergo post-translationally amidated at C-terminus. Each of these tridecapeptides possesses the most conserved and frequent N-terminus Asp-Ser-Ile (DSI). Sequence analysis revealed that Sj-PP/OK shared comparatively low identity with other invertebrates PP or OK. The tissue distribution profile showed differences in the expression level of Sj-PP/OK between male and female. qRT-PCR data demonstrated that Sj-PP/OK was widely distributed in various tissues, with its expression level increasing continuously in the brain, optic lobe, liver, and nidamental gland throughout the entire growth and development stages until gonad maturation. ISH detected that Sj-PP/OK positive signals existed in almost all regions of the optic lobe except the plexiform zone, the outer edge of all functional lobes in the brain, epithelial cells and the outer membrane layer of the accessory nidamental gland. These findings suggest that Sj-PP/OK might play a role in the regulation of reproduction, such as vitellogenin synthesis, restoration, and ova encapsulation. CONCLUSION: The study indicated that Sj-PP/OK may be involved in the neuroendocrine regulation in cephalopods, providing primary theoretical basis for further studies of its regulation role in reproduction.
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Secuencia de Aminoácidos , Decapodiformes , Hibridación in Situ , Neuropéptidos , Animales , Neuropéptidos/genética , Neuropéptidos/metabolismo , Neuropéptidos/química , Decapodiformes/genética , Decapodiformes/metabolismo , Hibridación in Situ/métodos , Filogenia , Sistemas de Lectura Abierta , Clonación Molecular , Secuencia de Bases , FemeninoRESUMEN
Insect neuropeptides play an essential role in regulating growth, development, reproduction, nerve conduction, metabolism, and behavior in insects; therefore, G protein-coupled receptors of neuropeptides are considered important targets for designing green insecticides. Cockroach-type allatostatins (ASTs) (FGLamides allatostatins) are important insect neuropeptides in Diploptera punctata that inhibit juvenile hormone (JH) synthesis in the corpora allata and affect growth, development, and reproduction of insects. Therefore, the pursuit of novel insecticides targeting the allatostatin receptor (AstR) holds significant importance. Previously, we identified an AST analogue, H17, as a promising candidate for pest control. Herein, we first modeled the 3D structure of AstR in D. punctata (Dippu-AstR) and predicted the binding mode of H17 with Dippu-AstR to study the critical interactions and residues favorable to its bioactivity. Based on this binding mode, we designed and synthesized a series of H17 derivatives and assessed their insecticidal activity against D. punctata. Among them, compound Q6 showed higher insecticidal activity than H17 against D. punctata by inhibiting JH biosynthesis, indicating that Q6 is a potential candidate for a novel insect growth regulator (IGR)-based insecticide. Moreover, Q6 exhibited insecticidal activity against Plutella xylostella, indicating that these AST analogs may have a wider insecticidal spectrum. The underlying mechanisms and molecular conformations mediating the interactions of Q6 with Dippu-AstR were explored to understand its effects on the bioactivity. The present work clarifies how a target-based strategy facilitates the discovery of new peptide mimics with better bioactivity, enabling improved IGR-based insecticide potency in sustainable agriculture.
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Proteínas de Insectos , Insecticidas , Neuropéptidos , Peptidomiméticos , Insecticidas/química , Insecticidas/farmacología , Insecticidas/síntesis química , Animales , Neuropéptidos/química , Neuropéptidos/farmacología , Neuropéptidos/metabolismo , Proteínas de Insectos/química , Proteínas de Insectos/metabolismo , Proteínas de Insectos/genética , Peptidomiméticos/química , Peptidomiméticos/farmacología , Peptidomiméticos/síntesis química , Diseño de Fármacos , Hormonas Juveniles/química , Hormonas Juveniles/farmacología , Hormonas Juveniles/metabolismo , Cucarachas/efectos de los fármacos , Cucarachas/químicaRESUMEN
Sunlight is part of our everyday life and most people accept it as beneficial to our health. With the advance of our knowledge in cutaneous photochemistry, photobiology and photomedicine over the past four decades, the terrestrial solar radiation has become a concern of dermatologists and is considered to be a major damaging environmental factor for our skin. Most photobiological effects (e.g., sunburn, suntanning, local and systemic immunosuppression, photoaging or dermatoheliosis, skin cancer and precancer, etc.) are attributed to ultraviolet radiation (UVR) and more particularly to UVB radiation (290-320 nm). UVA radiation (320-400 nm) also plays an important role in the induction of erythema by the photosensitized generation of reactive oxygen species (singlet oxygen ((1)O2))superoxide (O2-) and hydroxyl radicals ((OH) that damage DNA and cellular membranes, and promote carcinogenesis and the changes associated with photoaging. Therefore, research efforts have been directed at a better photochemical and photobiological understanding of the so-called sunburn reaction, actinic or solar erythema. To survive the insults of actinic damage, the skin appears to have different intrinsic defensive mechanisms, among which antioxidants (enzymatic and non-enzymatic systems) play a pivotal role. In this paper, we will review the basic aspects of the action of UVR on the skin: a) photochemical reactions resulting from photon absorption by endogenous chromophores; b) the lipid peroxidation phenomenon, and c) intrinsic defensive cutaneous mechanisms (antioxidant systems). The last section will cover the inflammatory response including mediator release after cutaneous UVR exposure and adhesion molecule expression.