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Without intervention, cardiac arrhythmias pose a risk of fatality. However, timely intervention can be challenging in environments where transporting a large, heavy defibrillator is impractical, or emergency surgery to implant cardiac stimulation devices is not feasible. Here, we introduce an injectable cardiac stimulator, a syringe loaded with a nanoparticle solution comprising a conductive polymer and a monomer that, upon injection, forms a conductive structure around the heart for cardiac stimulation. Following treatment, the electrode is cleared from the body, eliminating the need for surgical extraction. The mixture adheres to the beating heart in vivo without disrupting its normal rhythm. The electrofunctionalized injectable cardiac stimulator demonstrates a tissue-compatible Young's modulus of 21 kPa and a high conductivity of 55 S/cm. The injected electrode facilitates electrocardiogram measurements, regulates heartbeat in vivo, and rectifies arrhythmia. Conductive functionality is maintained for five consecutive days, and no toxicity is observed at the organism, organ, or cellular levels.
Asunto(s)
Arritmias Cardíacas , Animales , Arritmias Cardíacas/terapia , Arritmias Cardíacas/fisiopatología , Conductividad Eléctrica , Corazón/fisiología , Nanopartículas/química , Electrocardiografía , Humanos , Ratones , Frecuencia Cardíaca , Polímeros/química , Masculino , Inyecciones , Módulo de Elasticidad , Terapia por Estimulación Eléctrica/instrumentación , Terapia por Estimulación Eléctrica/métodos , Electrodos ImplantadosRESUMEN
Proteins are adjustable units from which biomaterials with designed properties can be developed. However, non-native folded states with controlled topologies are hardly accessible in aqueous environments, limiting their prospects as building blocks. Here, we demonstrate the ability of a series of anhydrous deep eutectic solvents (DESs) to precisely control the conformational landscape of proteins. We reveal that systematic variations in the chemical composition of binary and ternary DESs dictate the stabilization of a wide range of conformations, that is, compact globular folds, intermediate folding states, or unfolded chains, as well as controlling their collective behavior. Besides, different conformational states can be visited by simply adjusting the composition of ternary DESs, allowing for the refolding of unfolded states and vice versa. Notably, we show that these intermediates can trigger the formation of supramolecular gels, also known as eutectogels, where their mechanical properties correlate to the folding state of the protein. Given the inherent vulnerability of proteins outside the native fold in aqueous environments, our findings highlight DESs as tailorable solvents capable of stabilizing various non-native conformations on demand through solvent design.
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Geles , Pliegue de Proteína , Proteínas , Solventes , Solventes/química , Proteínas/química , Geles/química , Conformación ProteicaRESUMEN
Albumin-based cryogels for capturing haemin were synthesised by crosslinking different biomolecules, bovine serum albumin (BSA) and ovalbumin (OVA). The impact of the protein and coupling agent concentrations on cryogel's mechanical properties, swelling ratios and polymerisation yields, as well as autoclaving as a post-treatment on the cryogel, were studied. We found that BSA (50 mg/ml) and the crosslinker (N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride, 46 mg/ml) formed a cryogel with optimum physical characteristics at a comparatively low protein concentration. The cryogel's mechanical stability was increased using a double-layer cryogel approach by crosslinking the BSA proteins at subzero temperature inside an acrylamide and hydroxyethyl methacrylate premade cryogels. Batch binding and kinetic adsorption isotherms of haemin on the cryogels were assessed to evaluate their binding capacity toward the porphyrin molecule. The results showed that single-layer cryogels (BSA and OVA) had a higher capacity (â¼0.68 mg/ml gel) and higher reaction rate constant towards haemin adsorption than double-layer gels. In contrast, the double-layer cryogels had higher mechanical strength than single-layer gels. The experimental results suggested that the cryogels followed the Freundlich model and the pseudo-second-order isotherm for batch adsorption and kinetics, respectively. The interaction between haemin and the gels was studied by fluorescence quenching. We found between 1.1 and 1.6 binding sites for different cryogels.
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The preservation of labile biomolecules presents a major challenge in chemistry, and deep eutectic solvents (DESs) have emerged as suitable environments for this purpose. However, how the hydration of DESs impacts the behavior of proteins is often neglected. Here, we demonstrate that the amino acid environment and secondary structure of two proteins (bovine serum albumin and lysozyme) and an antibody (immunoglobulin G) in 1:2 choline chloride:glycerol and 1:2 choline chloride:urea follow a re-entrant behavior with solvent hydration. A dome-shaped transition is observed with a folded or partially folded structure at very low (<10 wt % H2O) and high (>40 wt % H2O) DES hydration, while protein unfolding increases between those regimes. Hydration also affects protein conformation and stability, as demonstrated for bovine serum albumin in hydrated 1:2 choline chloride:glycerol. In the neat DES, bovine serum albumin remains partially folded and unexpectedly undergoes unfolding and oligomerization at low water content. At intermediate hydration, the protein begins to refold and gradually retrieves the native monomer-dimer equilibrium. However, ca. 36 wt % H2O is required to recover the native folding fully. The half-denaturation temperature of the protein increases with decreasing hydration, but even the dilute DESs significantly enhance the thermal stability of bovine serum albumin. Also, protein unfolding can be reversed by rehydrating the sample to the high hydration regime, also recovering protein function. This correlation provides a new perspective to understanding protein behavior in hydrated DESs, where quantifying the DES hydration becomes imperative to identifying the folding and stability of proteins.
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Disolventes Eutécticos Profundos , Glicerol , Albúmina Sérica Bovina/química , Solventes/química , ColinaRESUMEN
Due to its compatible oxygen-transporting abilities, hemoglobin (Hb) is a protein of interest in the development of artificial oxygen therapeutics. Despite continuous formulation attempts, extracellular Hb solution often exhibits undesirable reactions when applied in vivo. Therefore, protein engineering is frequently used to examine alternative ways of controlling the unwanted reactions linked to cell-free Hb solutions. In this study, three mutants of human fetal hemoglobin (HbF) are evaluated; single mutants αA12D and αA19D, and a double mutant αA12D/A19D. These variants were obtained by site-directed mutagenesis and recombinant production in E. coli, and carry negative charges on the surface of the α-subunit at the designated mutation sites. Through characterization of the mutant proteins, we found that the substitutions affected the protein in several ways. As expected, the isoelectric points (pIs) were lowered, from 7.1 (wild-type) down to 6.6 (double mutant), which influenced the anion exchange chromatographic procedures by shifting conditions toward higher conductivity for protein elution. The biological and physiological properties of HbF could be improved by these small modifications on the protein surface. The DNA cleavage rate associated with native HbF could be reduced by 55%. In addition, the negatively charged HbF mutant had an extended circulation time when examined in a mouse model using top load Hb additions. At the same time, the mutations did not affect the overall structural integrity of the HbF molecule, as determined by small-angle X-ray scattering. In combination with circular dichroism and thermal stability, modest structural shifts imposed by the mutations could possibly be related to changes in secondary structure or reorganization. Such local deformations were too minor to be determined within the resolution of the structural data; and overall, unchanged oxidation and heme loss kinetics support the conclusion that the mutations did not adversely affect the basic structural properties of Hb. We confirm the value of adding negatively charged residues onto the surface of the protein to improve the global functions of recombinant Hb.
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How many nanoparticles can we load in a fiber? How much will leak? Underlying is the relatively new question of the "space available" in fibers for nanoparticle loading. Here, using supercritical carbon dioxide (scCO2) as a carrier fluid, we explored the impregnation in four Indian silks (Mulberry, Eri, Muga, and Tasar) with five standard sizes of gold nanoparticles (5, 20, 50, 100 and 150 nm in diameter). All silks could be permanently impregnated with nanoparticles up to 150 nm in size under scCO2 impregnation. Accompanying structural changes indicated that the amorphous silk domains reorganized to accommodate the gold NPs. The mechanism was studied in detail in degummed Mulberry silk fibers (i.e., without the sericin coating) with the 5 nm nanoparticle. The combined effects of concentration, time of impregnation, scCO2 pressure, and temperature showed that only a narrow set of conditions allowed for permanent impregnation without deterioration of the properties of the silk fibers.
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We have designed, built, and validated a (quasi)-simultaneous measurement platform called NUrF, which consists of neutron small-angle scattering, UV-visible, fluorescence, and densitometry techniques. In this contribution, we illustrate the concept and benefits of the NUrF setup combined with high-performance liquid chromatography pumps to automate the preparation and measurement of a mixture series of Brij35 nonionic surfactants with perfluorononanoic acid in the presence of a reporter fluorophore (pyrene).
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The foundations of silk spinning, the structure, storage, and activation of silk proteins, remain highly debated. By combining solution small-angle neutron and X-ray scattering (SANS and SAXS) alongside circular dichroism (CD), we reveal a shape anisotropy of the four principal native spider silk feedstocks from Nephila edulis. We show that these proteins behave in solution like elongated semiflexible polymers with locally rigid sections. We demonstrated that minor ampullate and cylindriform proteins adopt a monomeric conformation, while major ampullate and flagelliform proteins have a preference for dimerization. From an evolutionary perspective, we propose that such dimerization arose to help the processing of disordered silk proteins. Collectively, our results provide insights into the molecular-scale processing of silk, uncovering a degree of evolutionary convergence in protein structures and chemistry that supports the macroscale micellar/pseudo liquid crystalline spinning mechanisms proposed by the community.
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Seda , Arañas , Animales , Dicroismo Circular , Conformación Molecular , Dispersión del Ángulo Pequeño , Difracción de Rayos XRESUMEN
We have developed an in-situ method using sonication (3 mm probe sonicator, 30 W, 20 kHz) and auto-reduction (control) to study the mechanism of the formation of manganese dioxide (MnO2) on a solid template (silk film), and its resulting enzymatic activity on tetramethylbenzidine (TMB) substrate. The fabrication of the silk film was first optimized for stability (no degradation) and optical transparency. A factorial approach was used to assess the effect of sonication time and the initial concentration of potassium permanganate (KMnO4). The result indicated a significant correlation with a fraction of KMnO4 consumed and MnO2 formation. Further, we found that the optimal process conditions to obtain a stable silk film with highly catalytic MnO2 nanoparticles (NPs) was 30 min of sonication in the presence of 0.5 mM of KMnO4 at a temperature of 20-24 °C. Under the optimal condition, we monitored in-situ the formation of MnO2 on the silk film, and after thorough rinsing, the in-situ catalysis of 0.8 mM of TMB substrate. For control, we used the auto-reduction of KMnO4 onto the silk film after about 16 h. The result from single-wavelength analysis confirmed the different kinetics rates for the formation of MnO2 via sonication and auto-reduction. The result from the multivariate component analysis indicated a three components route for sonication and auto-reduction to form MnO2-Silk. Overall, we found that the smaller size, more mono-dispersed, and deeper buried MnO2 NPs in silk film prepared by sonication, conferred a higher catalytic activity and stability to the hybrid material.
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Materiales Biomiméticos/química , Enzimas/metabolismo , Compuestos de Manganeso/química , Nanopartículas/química , Óxidos/química , Seda/química , Sonicación , Bencidinas/química , Estabilidad de Medicamentos , Cinética , TemperaturaRESUMEN
A combination of supercritical carbon dioxide (scCO2) impregnation of pyrrole and sonochemical transformation of permanganate (KMnO4) was used to impart conductive and catalytic properties to silk fibers. The results indicated that the conductivity (from polypyrrole -PPy) and catalytic activities (from manganese dioxide -MnO2) were independent and complementary within the processing parameters used. The enhanced conductivity was attributed to scCO2 preferentially distributing the pyrrole monomers along with the silk internal fibrillar structure and hence, yielding a more linear PPy. The oxidative properties of the PPy-MnO2-silk hybrid showed an enzyme-like behavior for the degradation of hydrogen peroxide (H2O2) with a Km of about 13â¯mM and specific activity of 1470⯱â¯75⯵mol/min/g. Finally, we demonstrated that the PPy-MnO2-silk hybrid could be used as soft working electrodes for the simultaneous degradation and detection of H2O2.
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Técnicas Biosensibles , Conductividad Eléctrica , Peróxido de Hidrógeno/aislamiento & purificación , Seda/química , Dióxido de Carbono/química , Catálisis , Electrodos , Fibroínas/química , Peróxido de Hidrógeno/química , Compuestos de Manganeso/química , Óxidos/química , Polímeros/química , Pirroles/químicaRESUMEN
In the formulation of an active enzyme enclosed in a matrix for controlled delivery, it is a challenge to achieve a high protein load and to ensure high activity of the protein. For the first time to our knowledge, we report the use of a highly swollen lipid sponge (L3) phase for encapsulation of the large active enzyme, ß-galactosidase (ß-gal, 238 kDa). This enzyme has large relevance for applications in, e.g. the production of lactose free milk products. The formulation consisted of diglycerol monooleate (DGMO), and a mixture of mono-, di- and triglycerides (Capmul GMO-50) stabilised by polysorbate 80 (P80). The advantage of this type of matrix is that it can be produced on a large scale with a fairly simple and mild process as the system is in practice self-dispersing, yet it has a well-defined internal nano-structure. Minor effects on the sponge phase structure due to the inclusion of the enzyme were observed using small angle X-ray scattering (SAXS). The effect of encapsulation on the enzymatic activity and kinetic characteristics of ß-galactosidase activity was also investigated and can be related to the enzyme stability and confinement within the lipid matrix. The encapsulated ß-galactosidase maintained its activity for a significantly longer time when compared to the free solution at the same temperature. Differences in the particle size and charge of sponge-like nanoparticles (L3-NPs) with and without the enzyme were analysed by dynamic light scattering (DLS) and zeta-potential measurements. Moreover, all the initial ß-galactosidase was encapsulated within L3-NPs as revealed by size exclusion chromatography.
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Enzimas Inmovilizadas/química , Proteínas Fúngicas/química , Kluyveromyces/enzimología , Nanopartículas/química , beta-Galactosidasa/química , Caprilatos/química , Estabilidad de Enzimas , Glicéridos/química , Polisorbatos/química , Relación Estructura-ActividadRESUMEN
The protein Human Serum Albumin (HSA) is known to undergo conformational transitions towards partially unfolded forms triggered by acidification below pH 4.5. The extent of Fatty Acids (FA) binding has been thought to have an impact on the conformational equilibrium between the native and acid forms and to be a possible explanation for the observation of more than one band in early electrophoretic migration experiments at pH 4. We compared the acid-induced unfolding processes of commercial FA-free HSA, commercial "fatted" HSA and FA-HSA complexes, prepared at FA:HSA molar ratios between 1 and 6 by simple mixing and equilibration. We used a method for continuous acidification based on the hydrolysis of glucono-δ-lactone from pH 7 to pH 2.5, and followed the average protein changes by the blue shift of the intrinsic fluorescence emission and by performing a small angle X-ray scattering analysis on selected samples. The method also allowed for continuous monitoring of the increase of turbidity and laser light scattering of the protein samples related to the release of the insoluble ligands with acidification. Our results showed that the presence of FA interacting with albumin, an aspect often neglected in biophysical studies, affects the conformational response of the protein to acidification, and slightly shifts the loss of the native shape from pH 4.2 to pH 3.6. This effect increased with the FA:HSA molar ratio so that with three molar equivalents a saturation was reached, in agreement with the number of high-affinity binding sites reported for the FA. These findings confirm that a non-uniform level of ligand binding in an albumin sample can be an explanation for the early-observed conformational heterogeneity at pH 4.
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Ácidos/química , Ácidos Grasos/química , Conformación Proteica , Albúmina Sérica Humana/química , Ácidos Grasos/metabolismo , Humanos , Concentración de Iones de Hidrógeno , Isomerismo , Unión Proteica , Desplegamiento Proteico , Dispersión del Ángulo Pequeño , Albúmina Sérica Humana/metabolismo , Espectrometría de Fluorescencia , Difracción de Rayos XRESUMEN
The most abundant plasma protein, human serum albumin (HSA), is known to undergo several conformational transitions in an acidic environment. To avoid buffer effects and correlate global and local structural changes, we developed a continuous acidification method and simultaneously monitored the protein changes by both small-angle scattering (SAXS) and fluorescence. The progressive acidification, based on the hydrolysis of glucono-δ-lactone from pH 7 to pH 2.5, highlighted a multistep unfolding involving the putative F form (pH 4) and an extended and flexible conformation (pH < 3.5). The scattering profile of the F form was extracted by component analysis and further 3D modeled. The effect of acid unfolding at this intermediate stage was assigned to the rearrangement of the three albumin domains drifting apart toward a more elongated conformation, with a partial unfolding of one of the outer domains. To test the stabilizing effect of fatty acids, here palmitic acid, we compared the acid unfolding process of albumin with and without ligand. We found that when binding the ligand, the native conformation was favored up to lower pH values. Our approach solved the problem of realizing a continuous, homogeneous, and tunable acidification with simultaneous characterization applicable to study processes triggered by a pH decrease.
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Ácido Palmítico/química , Albúmina Sérica Humana/química , Sitios de Unión , Fluorescencia , Humanos , Concentración de Iones de Hidrógeno , Modelos Moleculares , Estabilidad Proteica , Desplegamiento Proteico , Dispersión del Ángulo Pequeño , Factores de Tiempo , Difracción de Rayos XRESUMEN
The most abundant plasma protein, human serum albumin (HSA), plays a key part in the body's antioxidant defense against reactive species. This study was aimed at correlating oxidant-induced chemical and structural effects on HSA. Despite the chemical modification induced by the oxidant hypochlorite, the native shape is preserved up to oxidant/HSA molar ratio <80, above which a structural transition occurs in the critical range 80-120. This conformational variation involves the drifting of one of the end-domains from the rest of the protein and corresponds to the loss of one-third of the α-helix and a net increase of the protein negative charge. The transition is highly reproducible suggesting that it represents a well-defined structural response typical of this multidomain protein. The ability to tolerate high levels of chemical modification in a folded or only partially unfolded state, as well as the stability to aggregation, provides albumin with optimal features as a biological buffer for the local formation of oxidants.
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Ácido Hipocloroso/síntesis química , Albúmina Sérica/química , Humanos , Ácido Hipocloroso/química , Ácido Hipocloroso/metabolismo , Oxidación-Reducción , Conformación Proteica , Albúmina Sérica/metabolismoRESUMEN
The fabrication of silk-based membranes that are stable, optically transparent and reusable is yet to be achieved. To address this bottleneck we have developed a method to produce transparent chromogenic silk patches that are optically responsive to pH. The patches were produced by blending regenerated silk fibroin (RSF), Laponite RD (nano clay) and the organic dyes neutral red and Thionine acetate. The Laponite RD played a central role in the patch mechanical integrity and prevention of dye leaching. The process was optimized using a factorial design to maximize the patch response to pH by UV absorbance and fluorescence emission. New patches of the optimized protocol, made from solutions containing 125 µM neutral red or 250 µM of Thionine and 15 mg/mL silk, were further tested for operational stability over several cycles of pH altering. Stability, performance, and reusability were achieved over the tested cycles. The approach could be extended to other reporting molecules or enzymes able to bind to Laponite.
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Colorantes/química , Tecnología de Fibra Óptica/instrumentación , Fibroínas/química , Rojo Neutro/química , Fenotiazinas/química , Silicatos/química , Animales , Bombyx/química , Bombyx/fisiología , Análisis Factorial , Fibroínas/aislamiento & purificación , Humanos , Concentración de Iones de Hidrógeno , Luz , Procesos FotoquímicosRESUMEN
The well-tuned spinning technology from spiders has attracted many researchers with the promise of producing high-performance, biocompatible, and yet biodegradable fibers. So far, the intricate chemistry and rheology of spinning have eluded us. A breakthrough was achieved recently, when the 3D structures of the N and C terminal domains of spider dragline silk were resolved and their pH-induced dimerization was revealed. To understand the terminal domains' dimerization mechanisms, we developed a protein model based on the experimental structures that reproduces charge and hydrophobic anisotropy of the complex protein surfaces. Monte Carlo simulations were used to study the thermodynamic dimerization of the N-terminal domain as a function of pH and ionic strength. We show that the hydrophobic and electrostatic anisotropies of the N-terminal domain cooperate constructively in the association process. The dipolar attractions at pH 6 lead to weakly bound dimers by forcing an antiparallel monomer orientation, stabilized by hydrophobic locking at close separations. Elevated salt concentrations reduce the thermodynamic dimerization constant due to screened electrostatic dipolar attraction. Moreover, the mutations on ionizable residues reveal a free energy of binding, proportional to the dipole moment of the mutants. It has previously been shown that dimers, formed at pH 6, completely dissociate at pH 7, which is thought to be due to altered protein charges. In contrast, our study indicates that the pH increase has no influence on the charge distribution of the N-terminal domain. Instead, the pH-induced dissociation is due to an adapted, loose conformation at pH 7, which significantly hampers both electrostatic and hydrophobic attractive interactions.
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In order to diagnose and fully correlate structural, chemical, and functional features of macromolecules and particles in solution, we propose the integration of spectroscopy and scattering on the same measuring volume and at the same time in a dedicated sample environment with multiple probes. Combined SAXS/UV-vis and SAXS/Raman information are employed to study the radiation damage effect in proteins in solution and the scattering from single wall carbon nanotubes (SWNTs) in SDS dispersion, respectively. In the first case, a clear correlation is observed between the time dependence of the radius of gyration (Rg) of the protein determined by SAXS and the turbidity of the protein solution extracted from simultaneous UV-vis measurements. In the second case, the ratio of bundled/isolated carbon nanotubes is obtained unambiguously through proper modeling of the scattering data and cross-validated with the Raman information. The uses of convex constraint analysis (CCA) and two-dimensional correlation analyses (2DCOS and 2DHCOS) are introduced to fully explore the combination of data sets from different techniques and to extract unique insights from the sample.
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Dispersión del Ángulo Pequeño , Espectrofotometría Ultravioleta/métodos , Espectrometría Raman/métodos , Difracción de Rayos X/métodos , Animales , Bovinos , Nanotubos de Carbono/química , Albúmina Sérica Bovina/química , Dodecil Sulfato de Sodio/químicaRESUMEN
Spider silk has been studied extensively for its attractive mechanical properties and potential applications in medicine and industry. The production of spider silk, however, has been lagging behind for lack of suitable systems. Our approach focuses on solving the production of spider silk by designing, expressing, purifying and characterizing the silk from cylindriform glands. We show that the cylindriform silk protein, in contrast to the commonly used dragline silk protein, is fully folded and stable in solution. With the help of GFP as a fusion tag we enhanced the expression of the silk protein in Escherichia coli and could optimize the downstream processing. Secondary structures analysis by circular dichroism and FTIR shows that the GFP-silk fusion protein is predominantly α-helical, and that pH can trigger a α- to ß-transition resulting in aggregation. Structural analysis by small angle X-ray scattering suggests that the GFP-Silk exists in the form of a hexamer in solution.
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Proteínas Fluorescentes Verdes/metabolismo , Multimerización de Proteína , Seda/química , Animales , Dicroismo Circular , Dispersión Dinámica de Luz , Electroforesis en Gel de Poliacrilamida , Fluorescencia , Concentración de Iones de Hidrógeno , Modelos Moleculares , Replegamiento Proteico , Estructura Secundaria de Proteína , Proteínas Recombinantes de Fusión/aislamiento & purificación , Dispersión del Ángulo Pequeño , Solubilidad , Espectroscopía Infrarroja por Transformada de Fourier , Arañas , Difracción de Rayos XRESUMEN
To better understand how silkmoth cocoons maintain the correct internal moisture levels for successful pupation, we examined cocoons from the long-domesticated mulberry silkmoth Bombyx mori as well as from two wild silkmoth species, Antheraea pernyi and Philosamia cynthia ricini. We determined fluid-independent values for the porosity, tortuosity and permeability of the inner and outer surfaces of cocoons. Permeabilities were low and, with the exception of A. pernyi cocoons, inner surfaces were less permeable than outer surfaces. B. mori cocoons exhibited the highest permeability overall, but only at the outer surface, while A. pernyi cocoons appeared to show different patterns from the other species tested. We discuss our findings in light of the ecophysiology of the various species and propose a 'tortuous path' model to help explain our results. The model describes how the structure of the inner and outer layers of the cocoon allows it to function as both a humidity trap and a waterproof barrier, providing optimum conditions for the successful development of the pupa.
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Bombyx/fisiología , Animales , Humedad , Mariposas Nocturnas/fisiología , Permeabilidad , Porosidad , Pupa/fisiologíaRESUMEN
This study investigates the relationship between birefringence and mechanical properties in the dragline silk of the gold orb weaving spider Nephila edulis. Using a custom birefringence-tensile testing device, we probed the orientation and water-induced swelling of fibers spun at variety of drawing rates ranging from 0.003 to 400 mm s(-1). Our results indicate that based upon drawing rate, silk fibers fall into three distinct regimes each with characteristic orientation and swelling properties. Further investigation using in situ tensile testing reveals interactions between a fiber's drawing speed, mechanical properties, and orientation that support previous model predictions. We propose that simultaneous birefringence-tensile testing provides a unique and readily accessible insight into the structural behavior of this interesting and important biomaterial.