RESUMEN
Mucin 7 (MUC7) is one of the salivary proteins whose role in the innate immune system is widely known, but still, neither its mechanism of action nor the impact of its metal coordination is fully understood. MUC7 and its fragments demonstrate potent antimicrobial activity, serving as a natural defense mechanism for organisms against pathogens. This study delves into the bioinorganic chemistry of MUC7 fragments (L1âEGRERDHELRHRRHHHQSPK; L2âEGRERDHELRHRR; L3âHHHQSPK) and their complexes with Cu(II) and Zn(II) ions. The antimicrobial characteristics of the investigated peptides and their complexes were systematically assessed against bacterial and fungal strains at pH 5.40 and pH 7.40. Our findings highlight the efficacy of these systems against Streptococcus sanguinis, a common oral cavity pathogen. Most interestingly, Zn(II) coordination increased (or triggered) the MUC7 antimicrobial activity, which underscores the pivotal role of metal ion coordination in governing the antimicrobial activity of human salivary MUC7 fragments against S. sanguinis.
Asunto(s)
Complejos de Coordinación , Cobre , Pruebas de Sensibilidad Microbiana , Mucinas , Proteínas y Péptidos Salivales , Zinc , Zinc/química , Zinc/farmacología , Humanos , Cobre/química , Cobre/farmacología , Complejos de Coordinación/farmacología , Complejos de Coordinación/química , Complejos de Coordinación/síntesis química , Mucinas/química , Mucinas/metabolismo , Mucinas/farmacología , Proteínas y Péptidos Salivales/farmacología , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/metabolismo , Antibacterianos/farmacología , Antibacterianos/química , Antibacterianos/síntesis química , Antifúngicos/farmacología , Antifúngicos/química , Antifúngicos/síntesis químicaRESUMEN
Environmental pollution with plastic polymers has become a global problem, leaving no continent and habitat unaffected. Plastic waste is broken down into smaller parts by environmental factors, which generate micro- and nanoplastic particles (MNPPs), ultimately ending up in the human food chain. Before entering the human body, MNPPs make their first contact with saliva in the human mouth. However, it is unknown what proteins attach to plastic particles and whether such protein corona formation is affected by the particle's biophysical properties. To this end, we employed polystyrene MNPPs of two different sizes and three different charges and incubated them individually with saliva donated by healthy human volunteers. Particle zeta potential and size analyses were performed using dynamic light scattering complemented by nanoliquid chromatography high-resolution mass spectrometry (nLC/HRMS) to qualitatively and quantitatively reveal the protein soft and hard corona for each particle type. Notably, protein profiles and relative quantities were dictated by plastic particle size and charge, which in turn affected their hydrodynamic size, polydispersity, and zeta potential. Strikingly, we provide evidence of the latter to be dynamic processes depending on exposure times. Smaller particles seemed to be more reactive with the surrounding proteins, and cultures of the particles with five different cell lines (HeLa, HEK293, A549, HepG2, and HaCaT) indicated protein corona effects on cellular metabolic activity and genotoxicity. In summary, our data suggest nanoplastic size and surface chemistry dictate the decoration by human saliva proteins, with important implications for MNPP uptake in humans.
Asunto(s)
Tamaño de la Partícula , Poliestirenos , Saliva , Proteínas y Péptidos Salivales , Propiedades de Superficie , Humanos , Saliva/química , Saliva/metabolismo , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/metabolismo , Poliestirenos/química , Corona de Proteínas/química , Corona de Proteínas/metabolismo , Nanopartículas/química , Microplásticos/químicaRESUMEN
Salivary complement inhibitors occur in many of the blood feeding arthropod species responsible for transmission of pathogens. During feeding, these inhibitors prevent the production of proinflammatory anaphylatoxins, which may interfere with feeding, and limit formation of the membrane attack complex which could damage arthropod gut tissues. Salivary inhibitors are, in many cases, novel proteins which may be pharmaceutically useful or display unusual mechanisms that could be exploited pharmaceutically. Albicin is a potent inhibitor of the alternative pathway of complement from the saliva of the malaria transmitting mosquito, Anopheles albimanus. Here we describe the cryo-EM structure of albicin bound to C3bBb, the alternative C3 convertase, a proteolytic complex that is responsible for cleavage of C3 and amplification of the complement response. Albicin is shown to induce dimerization of C3bBb, in a manner similar to the bacterial inhibitor SCIN, to form an inactive complex unable to bind the substrate C3. Size exclusion chromatography and structures determined after 30 minutes of incubation of C3b, factor B (FB), factor D (FD) and albicin indicate that FBb dissociates from the inhibited dimeric complex leaving a C3b-albicin dimeric complex which apparently decays more slowly.
Asunto(s)
Anopheles , Microscopía por Crioelectrón , Proteínas de Insectos , Animales , Proteínas de Insectos/química , Proteínas de Insectos/metabolismo , Anopheles/metabolismo , Anopheles/inmunología , Anopheles/parasitología , Complemento C3b/metabolismo , Complemento C3b/química , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/metabolismo , Modelos MolecularesRESUMEN
Ticks produce chemokine-binding proteins, known as evasins, in their saliva to subvert the host's immune response. Evasins bind to chemokines and thereby inhibit the activation of their cognate chemokine receptors, thus suppressing leukocyte recruitment and inflammation. We recently described subclass A3 evasins, which, like other class A evasins, exclusively target CC chemokines but appear to use a different binding site architecture to control target selectivity among CC chemokines. We now describe the structural basis of chemokine recognition by the class A3 evasin EVA-ACA1001. EVA-ACA1001 binds to almost all human CC chemokines and inhibits receptor activation. Truncation mutants of EVA-ACA1001 showed that, unlike class A1 evasins, both the N- and C-termini of EVA-ACA1001 play minimal roles in chemokine binding. To understand the structural basis of its broad chemokine recognition, we determined the crystal structure of EVA-ACA1001 in complex with the human chemokine CCL16. EVA-ACA1001 forms backbone-backbone interactions with the CC motif of CCL16, a conserved feature of all class A evasin-chemokine complexes. A hydrophobic pocket in EVA-ACA1001, formed by several aromatic side chains and the unique disulfide bond of class A3 evasins, accommodates the residue immediately following the CC motif (the "CC + 1 residue") of CCL16. This interaction is shared with EVA-AAM1001, the only other class A3 evasins characterized to date, suggesting it may represent a common mechanism that accounts for the broad recognition of CC chemokines by class A3 evasins.
Asunto(s)
Modelos Moleculares , Humanos , Animales , Garrapatas/química , Garrapatas/metabolismo , Cristalografía por Rayos X , Sitios de Unión , Proteínas de Artrópodos/química , Proteínas de Artrópodos/metabolismo , Proteínas de Artrópodos/genética , Unión Proteica , Quimiocinas/química , Quimiocinas/metabolismo , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/metabolismoRESUMEN
Using the surface characterization techniques of quartz crystal microbalance with dissipation, atomic force microscopy, and scanning electron microscopy, the structure of the salivary pellicle was investigated before and after it was exposed to dairy proteins, including micellar casein, skim milk, whey protein isolate (WPI), and a mixture of skim milk and WPI. We have shown that the hydration, viscoelasticity, and adsorbed proteinaceous mass of a preadsorbed salivary pellicle on a PDMS surface are greatly affected by the type of dairy protein. After interaction with whey protein, the preadsorbed saliva pellicle becomes softer. However, exposure of the saliva pellicle to micellar casein causes the pellicle to partially collapse, which results in a thinner and more rigid surface layer. This structure change correlates with the measured lubrication behavior when the saliva pellicle is exposed to dairy proteins. While previous studies suggest that whey protein is the main component in milk to interact with salivary proteins, our study indicates interactions with casein are more important. The knowledge gained here provides insights into the mechanisms by which different components of dairy foods and beverages contribute to mouthfeel and texture perception, as well as influence oral hygiene.
Asunto(s)
Película Dental , Proteínas y Péptidos Salivales , Película Dental/química , Película Dental/metabolismo , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/metabolismo , Adsorción , Caseínas/química , Caseínas/metabolismo , Propiedades de Superficie , Proteína de Suero de Leche/química , Humanos , Animales , Microscopía de Fuerza Atómica , Saliva/química , Saliva/metabolismo , Tecnicas de Microbalanza del Cristal de CuarzoRESUMEN
INTRODUCTION: Statherin-derived peptide (StatpSpS) has shown promise against erosive tooth wear. To elucidate its interaction with the hydroxyapatite (HAP) surface, the mechanism related to adsorption of this peptide with HAP was investigated through nanosecond-long all-atom molecular dynamics simulations. METHODS: StatpSpS was positioned parallel to the HAP surface in 2 orientations: 1 - neutral and negative residues facing the surface and 2 - positive residues facing the surface. A system containing StatpSpS without HAP was also simulated as control. In the case of systems with HAP, both partially restrained surface and unrestrained surface were constructed. Structural analysis, interaction pattern, and binding-free energy were calculated. RESULTS: In the peptide system without the HAP, there were some conformational changes during the simulation. In the presence of the surface, only moderate changes were observed. Many residues exhibited short and stable distances to the surface, indicating strong interaction. Specially, the residues ASP1 and SER2 have an important role to anchor the peptide to the surface, with positively charged residues, mainly arginine, playing a major role in the further stabilization of the peptide in an extended conformation, with close contacts to the HAP surface. CONCLUSION: The interaction between StatpSpS and HAP is strong, and the negative charged residues are important to the anchoring of the peptide in the surface, but after the initial placement the peptide rearranges itself to maximize the interactions between positive charged residues.
Asunto(s)
Durapatita , Simulación de Dinámica Molecular , Proteínas y Péptidos Salivales , Durapatita/química , Proteínas y Péptidos Salivales/química , Humanos , Adsorción , Propiedades de Superficie , Unión ProteicaRESUMEN
Aedes aegypti mosquitoes are important vectors of several debilitating and deadly arthropod-borne (arbo) viruses, including Yellow Fever virus, Dengue virus, West Nile virus and Zika virus (ZIKV). Arbovirus transmission occurs when an infected mosquito probes the host's skin in search of a blood meal. Salivary proteins from mosquitoes help to acquire blood and have also been shown to enhance pathogen transmission in vivo and in vitro. Here, we evaluated the interaction of mosquito salivary proteins with ZIKV by surface plasmon resonance and enzyme-linked immunosorbent assay. We found that three salivary proteins AAEL000793, AAEL007420, and AAEL006347 bind to the envelope protein of ZIKV with nanomolar affinities. Similar results were obtained using virus-like particles in binding assays. These interactions have no effect on viral replication in cultured endothelial cells and keratinocytes. Additionally, we found detectable antibody levels in ZIKV and DENV serum samples against the recombinant proteins that interact with ZIKV. These results highlight complex interactions between viruses, salivary proteins and antibodies that could be present during viral transmissions.
Asunto(s)
Aedes/metabolismo , Proteínas de Insectos/metabolismo , Mosquitos Vectores/metabolismo , Proteínas y Péptidos Salivales/metabolismo , Proteínas del Envoltorio Viral/metabolismo , Virus Zika/metabolismo , Aedes/química , Aedes/genética , Aedes/virología , Animales , Células Endoteliales/metabolismo , Células Endoteliales/virología , Proteínas de Insectos/química , Proteínas de Insectos/genética , Queratinocitos/metabolismo , Queratinocitos/virología , Cinética , Mosquitos Vectores/química , Mosquitos Vectores/genética , Mosquitos Vectores/virología , Unión Proteica , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/genética , Proteínas del Envoltorio Viral/química , Proteínas del Envoltorio Viral/genética , Replicación Viral , Virus Zika/química , Virus Zika/genéticaRESUMEN
BACKGROUND: Malaria is transmitted when infected Anopheles mosquitoes take a blood meal. During this process, the mosquitoes inject a cocktail of bioactive proteins that elicit antibody responses in humans and could be used as biomarkers of exposure to mosquito bites. This study evaluated the utility of IgG responses to members of the Anopheles gambiae D7 protein family as serological markers of human-vector contact. METHODS: The D7L2, D7r1, D7r2, D7r3, D7r4 and SG6 salivary proteins from An. gambiae were expressed as recombinant antigens in Escherichia coli. Antibody responses to the salivary proteins were compared in Europeans with no prior exposure to malaria and lifelong residents of Junju in Kenya and Kitgum in Uganda where the intensity of malaria transmission is moderate and high, respectively. In addition, to evaluate the feasibility of using anti-D7 IgG responses as a tool to evaluate the impact of vector control interventions, we compared responses between individuals using insecticide-treated bednets to those who did not in Junju, Kenya where bednet data were available. RESULTS: We show that both the long and short forms of the D7 salivary gland antigens elicit a strong antibody response in humans. IgG responses against the D7 antigens reflected the transmission intensities of the three study areas, with the highest to lowest responses observed in Kitgum (northern Uganda), Junju (Kenya) and malaria-naïve Europeans, respectively. Specifically, the long form D7L2 induced an IgG antibody response that increased with age and that was lower in individuals who slept under a bednet, indicating its potential as a serological tool for estimating human-vector contact and monitoring the effectiveness of vector control interventions. CONCLUSIONS: This study reveals that D7L2 salivary antigen has great potential as a biomarker of exposure to mosquito bites and as a tool for assessing the efficacy of vector control strategies such as bednet use.
Asunto(s)
Anopheles/química , Mordeduras y Picaduras de Insectos/epidemiología , Proteínas del Tejido Nervioso/inmunología , Proteínas y Péptidos Salivales/química , Adolescente , Animales , Anopheles/fisiología , Biomarcadores/química , Niño , Preescolar , Estudios de Cohortes , Estudios Transversales , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Inmunoglobulina G/sangre , Lactante , Mordeduras y Picaduras de Insectos/diagnóstico , Kenia , Proteínas del Tejido Nervioso/química , Proteínas y Péptidos Salivales/inmunologíaRESUMEN
Erectile dysfunction (ED) is one of the most common sexual disorders in men. During the past 30 years, there has been no new drug development for ED. Thus, exploring the genetic basis of ED deserves further study, in hope of developing new pharmacological treatments for ED. In this study, Real-Time PCR analysis was used to assess the expression of androgen regulatory protein (Andpro) and pyruvate dehydrogenase kinase 4 (Pdk4) genes in ED. For this purpose, the experiment was performed on 20 men with severe ED and 20 potent men. IIEF-15 was used to determine the ED severity. The study was conducted in the Department of Sexual Medicine of the Kermanshah University of Medical Sciences, Kermanshah, Iran. The EDTA-Na vacuum blood tube was taken from ED patients and controls. Informed consent was obtained from all participants. After blood sampling, RNA was extracted from whole blood. Then cDNA was synthesized. The gene expression was analyzed through the qPCR method. The ß-actin was used as a reference gene. To further study these two proteins, their three-dimensional structures were predicted through I-TASSER. Compared with controls, in ED patients, the expression of the Andpro gene decreased, while the expression of the Pdk4 gene increased (p<0.01). Predicting the structure of the protein showed that Pyruvate Dehydrogenase Kinase 4 had a double subunit and androgen-regulated protein had a single subunit.
Asunto(s)
Disfunción Eréctil , Regulación de la Expresión Génica , Proteínas Quinasas , Proteínas y Péptidos Salivales , Adulto , Humanos , Masculino , Persona de Mediana Edad , Algoritmos , Secuencia de Aminoácidos , Biología Computacional/métodos , Disfunción Eréctil/genética , Disfunción Eréctil/metabolismo , Disfunción Eréctil/patología , Modelos Moleculares , Conformación Proteica , Proteínas Quinasas/química , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/genética , Proteínas y Péptidos Salivales/metabolismo , Índice de Severidad de la EnfermedadRESUMEN
The salivary glands of the flea Xenopsylla cheopis, a vector of the plague bacterium, Yersinia pestis, express proteins and peptides thought to target the hemostatic and inflammatory systems of its mammalian hosts. Past transcriptomic analyses of salivary gland tissue revealed the presence of two similar peptides (XC-42 and XC-43) having no extensive similarities to any other deposited sequences. Here we show that these peptides specifically inhibit coagulation of plasma and the amidolytic activity of α-thrombin. XC-43, the smaller of the two peptides, is a fast, tight-binding inhibitor of thrombin with a dissociation constant of less than 10 pM. XC-42 exhibits similar selectivity as well as kinetic and binding properties. The crystal structure of XC-43 in complex with thrombin shows that despite its substrate-like binding mode, XC-43 is not detectably cleaved by thrombin and that it interacts with the thrombin surface from the enzyme catalytic site through the fibrinogen-binding exosite I. The low rate of hydrolysis was verified in solution experiments with XC-43, which show the substrate to be largely intact after 2 h of incubation with thrombin at 37 °C. The low rate of XC-43 cleavage by thrombin may be attributable to specific changes in the catalytic triad observable in the crystal structure of the complex or to extensive interactions in the prime sites that may stabilize the binding of cleavage products. Based on the increased arterial occlusion time, tail bleeding time, and blood coagulation parameters in rat models of thrombosis XC-43 could be valuable as an anticoagulant.
Asunto(s)
Anticoagulantes/química , Antitrombinas/química , Proteínas de Insectos/química , Glándulas Salivales/química , Proteínas y Péptidos Salivales/química , Trombina , Xenopsylla/química , Animales , Humanos , Ratas , Trombina/antagonistas & inhibidores , Trombina/química , Xenopsylla/metabolismoRESUMEN
The sialotranscriptomes of Aedes aegypti revealed a transcript overexpressed in female salivary glands that codes a mature 7.8 kDa peptide. The peptide, specific to the Aedes genus, has a unique sequence, presents a putative secretory nature and its function is unknown. Here, we confirmed that the peptide is highly expressed in the salivary glands of female mosquitoes when compared to the salivary glands of males, and its secretion in mosquito saliva is able to sensitize the vertebrate host by inducing the production of specific antibodies. The synthetic version of the peptide downmodulated nitric oxide production by activated peritoneal murine macrophages. The fractionation of a Ae. aegypti salivary preparation revealed that the fractions containing the naturally secreted peptide reproduced the nitric oxide downmodulation. The synthetic peptide also selectively interfered with cytokine production by murine macrophages, inhibiting the production of IL-6, IL-12p40 and CCL2 without affecting TNF-α or IL-10 production. Likewise, intracellular proteins associated with macrophage activation were also distinctively modulated: while iNOS and NF-κB p65 expression were diminished, IκBα and p38 MAPK expression did not change in the presence of the peptide. The anti-inflammatory properties of the synthetic peptide were tested in vivo on a dextran sulfate sodium-induced colitis model. The therapeutic administration of the Ae. aegypti peptide reduced the leukocytosis, macrophage activity and nitric oxide levels in the gut, as well as the expression of cytokines associated with the disease, resulting in amelioration of its clinical signs. Given its biological properties in vitro and in vivo, the molecule was termed Aedes-specific MOdulatory PEptide (AeMOPE-1). Thus, AeMOPE-1 is a novel mosquito-derived immunobiologic with potential to treat immune-mediated disorders.
Asunto(s)
Aedes/inmunología , Colitis/etiología , Colitis/metabolismo , Activación de Macrófagos/inmunología , Macrófagos/inmunología , Proteínas y Péptidos Salivales/inmunología , Secuencia de Aminoácidos , Animales , Biomarcadores , Colitis/patología , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Femenino , Inmunomodulación , Activación de Linfocitos/inmunología , Macrófagos/metabolismo , Masculino , Ratones , Proteínas y Péptidos Salivales/química , Linfocitos T/inmunología , Linfocitos T/metabolismoRESUMEN
The aim of this study was to investigate the atomic composition and the proteome of the salivary proteins adsorbed on the surface of orthodontic metallic bracket. For this, the atomic composition of orthodontic metallic brackets was analyzed with X-ray Photoelectron Spectroscopy (XPS). The acquired bracket pellicle was characterized after brackets were immersed in human whole saliva supernatant for 2 hours at 37°C. Hydroxyapatite (HA) discs were used as a control. Acquired pellicle was harvested from the HA discs (n = 12) and from the metallic brackets (n = 12). Proteomics based on mass spectrometry technology was used for salivary protein identification and characterization. Results showed that most of the proteins adsorbed on the surface of orthodontic metallic brackets and on the HA discs were identified specifically to each group, indicating a small overlapping between the salivary proteins on each study group. A total of 311 proteins present on the HA discs were unique to this group while 253 proteins were unique to metallic brackets, and only 45 proteins were common to the two groups. Even though most proteins were unique to each study group, proteins related to antimicrobial activity, lubrication, and remineralization were present in both groups. These findings demonstrate that the salivary proteins adsorbed on the bracket surface are dependent on the material molecular composition.
Asunto(s)
Durapatita/química , Soportes Ortodóncicos , Proteoma/análisis , Proteínas y Péptidos Salivales/química , Adsorción , Cromatografía Líquida de Alta Presión , Humanos , Metales/química , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
During feeding, a tick's mouthpart penetrates the host's skin and damages tissues and small blood vessels, triggering the extrinsic coagulation and lectin complement pathways. To elude these defense mechanisms, ticks secrete multiple anticoagulant proteins and complement system inhibitors in their saliva. Here, we characterized the inhibitory activities of the homologous tick salivary proteins tick salivary lectin pathway inhibitor, Salp14, and Salp9Pac from Ixodesscapularis in the coagulation cascade and the lectin complement pathway. All three proteins inhibited binding of mannan-binding lectin to the polysaccharide mannan, preventing the activation of the lectin complement pathway. In contrast, only Salp14 showed an appreciable effect on coagulation by prolonging the lag time of thrombin generation. We found that the anticoagulant properties of Salp14 are governed by its basic tail region, which resembles the C terminus of tissue factor pathway inhibitor alpha and blocks the assembly and/or activity of the prothrombinase complex in the same way. Moreover, the Salp14 protein tail contributes to the inhibition of the lectin complement pathway via interaction with mannan binding lectin-associated serine proteases. Furthermore, we identified BaSO4-adsorbing protein 1 isolated from the tick Ornithodoros savignyi as a distant homolog of tick salivary lectin pathway inhibitor/Salp14 proteins and showed that it inhibits the lectin complement pathway but not coagulation. The structure of BaSO4-adsorbing protein 1, solved here using NMR spectroscopy, indicated that this protein adopts a noncanonical epidermal growth factor domain-like structural fold, the first such report for tick salivary proteins. These data support a mechanism by which tick saliva proteins simultaneously inhibit both the host coagulation cascade and the lectin complement pathway.
Asunto(s)
Proteínas de Artrópodos/ultraestructura , Interacciones Huésped-Patógeno/genética , Lectinas/genética , Proteínas y Péptidos Salivales/ultraestructura , Animales , Proteínas de Artrópodos/química , Proteínas de Artrópodos/genética , Coagulación Sanguínea/genética , Vasos Sanguíneos/parasitología , Vasos Sanguíneos/patología , Lectina de Unión a Manosa de la Vía del Complemento/genética , Ixodes/patogenicidad , Ixodes/ultraestructura , Lectinas/ultraestructura , Espectroscopía de Resonancia Magnética , Conformación Proteica , Saliva/química , Saliva/metabolismo , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/genética , Trombina/genética , Garrapatas/genética , Garrapatas/patogenicidadRESUMEN
Polyphenols have been part of human culture for about 6000 years. However, their mode of action in relation to wine tasting while eating is only beginning to be understood. This review, using analytical techniques and physicochemical concepts, attempts to summarize current knowledge and present an integrated view of the complex relationship between tannins, salivary proteins, lipids in food and in oral membranes. The action of tannins on taste sensations and astringency depends on their colloidal state. Although taste sensations are most likely due to interactions with taste receptors, astringency results from strong binding to proline-rich salivary proteins that otherwise lubricate the palate. Tannins disorder non-keratinized mucosa in mouth, possibly perturbing taste receptor function. The 10-15% ethanol present in wines potentiates this action. Cholesterol present in large quantities in keratinized mucosa prevents any disordering action on these oral membranes. Polyphenols bind strongly to the lipid droplets of fatty foods, a situation that reduces the astringency perceived when drinking a tannic wine, the so-called "camembert effect". Based on binding constants mainly measured by NMR, a comprehensive thermodynamic model of the interrelation between polyphenols, salivary proteins, lipids and taste receptors is presented.
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Lípidos de la Membrana/química , Proteínas y Péptidos Salivales/química , Taninos/química , Vino/análisis , Coloides/química , Etanol/análisis , Humanos , Micelas , GustoRESUMEN
The saliva of blood-sucking leeches contains a plethora of anticoagulant substances. One of these compounds derived from Haementeria ghilianii, the 66mer three-disulfide-bonded peptide tridegin, specifically inhibits the blood coagulation factor FXIIIa. Tridegin represents a potential tool for antithrombotic and thrombolytic therapy. We recently synthesized two-disulfide-bonded tridegin variants, which retained their inhibitory potential. For further lead optimization, however, structure information is required. We thus analyzed the structure of a two-disulfide-bonded tridegin isomer by solution 2D NMR spectroscopy in a combinatory approach with subsequent MD simulations. The isomer was studied using two fragments, i.e., the disulfide-bonded N-terminal (Lys1-Cys37) and the flexible C-terminal part (Arg38-Glu66), which allowed for a simplified, label-free NMR-structure elucidation of the 66mer peptide. The structural information was subsequently used in molecular modeling and docking studies to provide insights into the structure-activity relationships. The present study will prospectively support the development of anticoagulant-therapy-relevant compounds targeting FXIIIa.
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Factor XIIIa/antagonistas & inhibidores , Espectroscopía de Resonancia Magnética/métodos , Proteínas y Péptidos Salivales/farmacología , Secuencia de Aminoácidos , Animales , Disulfuros/química , Factor XIIIa/metabolismo , Fibrinolíticos/farmacología , Humanos , Isomerismo , Sanguijuelas/metabolismo , Imagen por Resonancia Magnética/métodos , Modelos Moleculares , Simulación de Dinámica Molecular , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/metabolismo , Relación Estructura-ActividadRESUMEN
BACKGROUND: The invasion of the mosquito salivary glands by Plasmodium sporozoites is a critical step that defines the success of malaria transmission and a detailed understanding of the molecules responsible for salivary gland invasion could be leveraged towards control of vector-borne pathogens. Antibodies directed against the mosquito salivary gland protein SGS1 have been shown to reduce Plasmodium gallinaceum sporozoite invasion of Aedes aegypti salivary glands, but the specific role of this protein in sporozoite invasion and in other stages of the Plasmodium life cycle remains unknown. METHODS: RNA interference and CRISPR/Cas9 were used to evaluate the role of A. aegypti SGS1 in the P. gallinaceum life cycle. RESULTS: Knockdown and knockout of SGS1 disrupted sporozoite invasion of the salivary gland. Interestingly, mosquitoes lacking SGS1 also displayed fewer oocysts. Proteomic analyses confirmed the abolishment of SGS1 in the salivary gland of SGS1 knockout mosquitoes and revealed that the C-terminus of the protein is absent in the salivary gland of control mosquitoes. In silico analyses indicated that SGS1 contains two potential internal cleavage sites and thus might generate three proteins. CONCLUSION: SGS1 facilitates, but is not essential for, invasion of A. aegypti salivary glands by P. gallinaceum and has a dual role as a facilitator of parasite development in the mosquito midgut. SGS1 could, therefore, be part of a strategy to decrease malaria transmission by the mosquito vector, for example in a transgenic mosquito that blocks its interaction with the parasite.
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Aedes/genética , Proteínas de Insectos/genética , Plasmodium gallinaceum/fisiología , Proteínas y Péptidos Salivales/genética , Aedes/parasitología , Secuencia de Aminoácidos , Animales , Femenino , Tracto Gastrointestinal/parasitología , Proteínas de Insectos/química , Proteínas de Insectos/metabolismo , Mosquitos Vectores/genética , Mosquitos Vectores/parasitología , Glándulas Salivales/parasitología , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/metabolismo , Alineación de Secuencia , Esporozoítos/fisiologíaRESUMEN
Mucin 7 (encoded byMUC7) is a human salivary protein that has a role in the natural immune system. Fragments of mucin 7 exhibit antimicrobial activity against bacteria and yeast. Although the antimicrobial properties of peptides have been known and studied for decades, the exact mechanism of action of antimicrobial peptides (AMPs) is still unclear. It is known that some AMPs require divalent metal ions to activate their activity. Herein, we investigated three 15-mer MUC7 peptides, one of which (mother peptide, sequence, L3) is a synthetic analog of a fragment naturally excised from MUC7 (with His3, His8, and His 14) and its two structural analogs, containing only two histidine residues, His3, His13 and His8, His13 (L2 and L1, respectively). Since there is a correlation between lipophilicity, the presence of metal ions (such as Cu(II) and Zn(II)) and antimicrobial activity of AMP, antimicrobial properties of the studied peptides, as well as their complexes with Cu(II) and Zn(II) ions, were tested for activity against Gram-positive (Enterococcus faecalis, Staphylococcus epidermidis) and Gram-negative (Escherichia coli, Pseudomonas aeruginosa) bacteria and fungi (Candida albicans). The results were correlated with their lipophilicity. Coordination and thermodynamic studies (potentiometry, UV-Vis, CD) revealed the formation of mainly mononuclear complexes in solution for all studied systems with different stability in the physiological pH range.
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Péptidos Antimicrobianos/farmacología , Complejos de Coordinación/farmacología , Mucinas/farmacología , Proteínas y Péptidos Salivales/farmacología , Secuencia de Aminoácidos , Péptidos Antimicrobianos/química , Humanos , Pruebas de Sensibilidad Microbiana , Mucinas/química , Proteínas y Péptidos Salivales/química , TermodinámicaRESUMEN
Aedes aegypti saliva facilitates blood meal acquisition through pharmacologically active compounds that prevent host hemostasis. Among these salivary proteins are the D7s, which are highly abundant and have been shown to act as scavengers of biogenic amines and eicosanoids. In this work, we performed comparative structural modeling, characterized the binding capabilities, and assessed the physiological functions of the Ae. aegypti salivary protein AeD7L2 compared to the well-characterized AeD7L1. AeD7L1 and AeD7L2 show different binding affinities to several biogenic amines and biolipids involved in host hemostasis. Interestingly, AeD7L2 tightly binds U-46619, the stable analog of thromboxane A2 (KD = 69.4 nm), which is an important platelet aggregation mediator, while AeD7L1 shows no binding. We tested the ability of these proteins to interfere with the three branches of hemostasis: vasoconstriction, platelet aggregation, and blood coagulation. Pressure myography experiments showed these two proteins reversed isolated resistance artery vasoconstriction induced by either norepinephrine or U-46619. These proteins also inhibited platelet aggregation induced by low doses of collagen or U-46619. However, D7 long proteins did not affect blood coagulation. The different ligand specificity and affinities of AeD7L1 and AeD7L2 matched our experimental observations from studying their effects on vasoconstriction and platelet aggregation, which confirm their role in preventing host hemostasis. This work highlights the complex yet highly specific biological activities of mosquito salivary proteins and serves as another example of the sophisticated biology underlying arthropod blood feeding.
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Aedes/metabolismo , Dengue/metabolismo , Proteínas de Insectos/metabolismo , Mosquitos Vectores/metabolismo , Proteínas y Péptidos Salivales/metabolismo , Ácido 15-Hidroxi-11 alfa,9 alfa-(epoximetano)prosta-5,13-dienoico/metabolismo , Aedes/fisiología , Aedes/virología , Secuencia de Aminoácidos , Animales , Coagulación Sanguínea/fisiología , Dengue/virología , Virus del Dengue/fisiología , Conducta Alimentaria/fisiología , Expresión Génica , Proteínas de Insectos/química , Proteínas de Insectos/genética , Modelos Moleculares , Mosquitos Vectores/virología , Agregación Plaquetaria/fisiología , Unión Proteica , Conformación Proteica , Proteínas y Péptidos Salivales/química , Proteínas y Péptidos Salivales/genética , Homología de Secuencia de AminoácidoRESUMEN
Astringency is an important quality attribute of green tea infusion, and (-)-Epigallocatechin gallate (EGCG) is the main contributor to astringency. Turbidity was used to predict the intensity of astringency for EGCG. The interactions between the selected proteins and EGCG, and the impacts of temperature, pH, protein structure, and EGCG concentration were studied. Mucin was selected as the protein in study for the prediction of EGCG astringency intensity. A predictive model (R2 = 0.994) was developed based on the relationship between the astringency of EGCG and the turbidity of EGCG/mucin mixtures at pH 5.0 and 37 °C. The fluorescence quenching analyses showed the interactions between EGCG and the selected proteins, which induced the reversible protein molecule conformational changes. The interactions were considered as the main reason that causes the astringency of tea infusions. The results provided a biochemical approach to explore the sensory qualities of green tea.
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Catequina/análogos & derivados , Proteínas y Péptidos Salivales/química , Té/química , Adulto , Catequina/química , Femenino , Fluorescencia , Humanos , Concentración de Iones de Hidrógeno , Masculino , Persona de Mediana Edad , Modelos Teóricos , Mucinas/química , Conformación Proteica , Espectrometría de Fluorescencia , Gusto , TemperaturaRESUMEN
MUC5B and MUC7 salivary mucins are reported to inhibit HIV-1 entry into target cells in vitro; however, their relative inhibitory potencies have not been quantitively compared. There is also conflicting evidence regarding whether HIV-1 infection diminishes mucins' inhibitory efficacy. We explored the effect of donor HIV-1 status upon the anti-HIV-1 potency of purified MUC5B and MUC7 while comparing their relative inhibitory potential using a pseudovirus-based neutralization assay. HIV status of sample donors had no detectable effect on HIV-1 inhibition by salivary mucins. MUC5B (median IC50 50 µg/ml, IQR 10-116 µg/ml) exhibited significantly more potent HIV-1 inhibition than MUC7 (median IC50 458 µg/ml, IQR 192->2000 µg/ml; Mann-Whitney U p < 0.0001). We suggest that larger size, gel-forming properties and extensive glycosylation of MUC5B allow more effective binding and aggregation of viral particles. MUC5B is also more abundant in the saliva and is therefore likely to make a substantially greater contribution to it's anti-HIV-1 properties.