| 1. |
- Clifton, L. A., et al.
(författare)
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Characterizing biomaterial complexity
- 2009
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Ingår i: Materials Today. - 1369-7021. ; 12:7-8, s. 86-91
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Tidskriftsartikel (refereegranskat)abstract
- Biomaterials research will always require a range of techniques to examine structure and function on a range of length scales and in a range of settings. Neutron scattering provides a unique way of disentangling the molecular and structural complexity of biomaterials through study of the constituent components. We examine how the technique has been used to study surface immobilized proteins and lipid films, floating lipid bilayers as mimics of in vitro planar membranes, and formation of fibres from solution by insects and spiders.
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| 2. |
- Del Giudice, Alessandra, et al.
(författare)
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Structural response of human serum albumin to oxidation : Biological buffer to local formation of hypochlorite
- 2016
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Ingår i: Journal of Physical Chemistry B. - : American Chemical Society (ACS). - 1520-6106 .- 1520-5207. ; 120:40, s. 12261-12271
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Tidskriftsartikel (refereegranskat)abstract
- 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. (Graph Presented).
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| 3. |
- Del Giudice, Alessandra, et al.
(författare)
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The effect of fatty acid binding in the acid isomerizations of albumin investigated with a continuous acidification method
- 2018
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Ingår i: Colloids and Surfaces B: Biointerfaces. - : Elsevier BV. - 0927-7765. ; 168, s. 109-116
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Tidskriftsartikel (refereegranskat)abstract
- 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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| 4. |
- Del Giudice, Alessandra, et al.
(författare)
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Time-Dependent pH Scanning of the Acid-Induced Unfolding of Human Serum Albumin Reveals Stabilization of the Native Form by Palmitic Acid Binding
- 2017
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Ingår i: Journal of Physical Chemistry B. - : American Chemical Society (ACS). - 1520-6106 .- 1520-5207. ; 121:17, s. 4388-4399
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Tidskriftsartikel (refereegranskat)abstract
- 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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| 5. |
- Dicko, Cedric, et al.
(författare)
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Differential scanning fluorimetry illuminates silk feedstock stability and processability.
- 2016
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Ingår i: Soft Matter. - : Royal Society of Chemistry (RSC). - 1744-6848 .- 1744-683X. ; 12:1, s. 255-262
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Tidskriftsartikel (refereegranskat)abstract
- The ability to design and implement silk feedstock formulations for tailored spinning has so far eluded the bioengineers. Recently, the high throughput screening technique of differential scanning fluorimetry (DSF) demonstrated the link between the instability transition temperature (Ti) and the processability of the silk feedstock. Using DSF we screened a large set of chemicals known to affect solvent quality. A multivariate analysis of the results shows that, regardless of the diversity of chemicals, three groupings are significantly distinguishable: G1 = similar to native silk; G2 = largely dominated by electrostatic interactions; and G3 = dominated by chelating interactions. We propose a thermodynamic analysis based on a pre- and post-transition fit to estimate the van't Hoff enthalpies (ΔHv) and the instability temperature (Ti). Our analysis shows that the ΔTi and ΔHv values were distinct: G1 (ΔTi = 0.23 ± 0.2; ΔHv = -159.1 ± 5.6 kcal mol(-1)), G2 (ΔTi = -7.3 ± 0.7; ΔHv = -191.4 ± 5.5 kcal mol(-1)), and G3 (ΔTi = -19.9 ± 3.3; ΔHv = -68.8 ± 6.0 kcal mol(-1)). Our analysis further combined the ΔTi value and the ΔHv value using stability ΔΔG to find that G1 only marginally stabilizes native silks (ΔΔG = -0.15 ± 0.04 kcal mol(-1)), whereas G2 and G3 destabilize native silk (ΔΔG = 3.8 ± 0.11 and ΔΔG = 3.8 ± 0.3 kcal mol(-1), respectively). Here our analysis shows that native silk has a complex multistep transition that is possibly non-cooperative. However, all three groupings also show a direct and cooperative transition with varied stabilization effects. This analysis suggests that native silks are able to sample multiple substates prior to undergoing (or to delay) the final transition. We conclude by hypothesizing that the observed energetic plasticity may be mediated by a fragile packaging of the silk tertiary structure that is readily lost when the solvent quality changes.
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| 6. |
- Dicko, Cedric, et al.
(författare)
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NUrF-Optimization of in situ UV-vis and fluorescence and autonomous characterization techniques with small-angle neutron scattering instrumentation
- 2020
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Ingår i: Review of Scientific Instruments. - : AMER INST PHYSICS. - 0034-6748 .- 1089-7623. ; 91:7
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Tidskriftsartikel (refereegranskat)abstract
- 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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| 7. |
- Ferreira, Betina M.P., et al.
(författare)
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3D Structure and Mechanics of Silk Sponge Scaffolds Is Governed by Larger Pore Sizes
- 2020
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Ingår i: Frontiers in Materials. - : Frontiers Media SA. - 2296-8016. ; 7
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Tidskriftsartikel (refereegranskat)abstract
- Three-dimensional scaffolds play an essential role in tissue engineering. Although essential, the tunability of the 3D scaffolds mechanical and transport properties remains a challenge. In this work, we present new approaches to advance the field. First, we applied our progressive pH acidification to mimic the natural silk gelation process before ice-templating (−20 and −80°C); second, we fitted the mechanical properties using a connectivity model; third, we fitted the scaffolds mechanical relaxation to understand the transport properties; and fourth we used micro-CT to correlate the process parameters to the scaffolds' performances. Our results suggested that the free shrinkage of the scaffolds determined their final properties. We found, however, that the porosity (above 90%) was anisotropic, similarly the tortuosity (between 1 and 1.3). We identified two major pore dimensions, the first one between 10 and 20 μm, and the second between 50 and 130 μm. Mechanically, our model suggested that the bulk modulus captured the elastic contribution and was controlled predominantly by the silk concentration. We tentatively associated the fractional modulus 1 to the collapse of the larger pores structures and was controlled mostly by the process temperature. We assigned the slow relaxation to the transport of fluid in the silk sponge scaffolds; and the fast relaxation with a viscoelastic relaxation. The silk concentration and process temperatures did not influence the latter. Overall, our use of the tomography, mechanical test, and detailed statistical analysis provides inroads into the interplay between process parameters (silk concentration and process temperature) and the multiple responses of the silk sponge scaffolds. The development of a new mechanical fitting for the compression test helped capture simply the different failure modes in the sponge scaffolds as well as correlating those events to relaxation and eventually transport properties.
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| 8. |
- Gilbert, Jennifer, et al.
(författare)
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Immobilisation of β-galactosidase within a lipid sponge phase: structure, stability and kinetics characterisation
- 2019
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Ingår i: Nanoscale. - 2040-3372. ; 11:44, s. 21291-21301
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Tidskriftsartikel (refereegranskat)abstract
- 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, β-galactosidase (β-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 β-galactosidase activity was also investigated and can be related to the enzyme stability and confinement within the lipid matrix. The encapsulated β-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 β-galactosidase was encapsulated within L3-NPs as revealed by size exclusion chromatography.
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| 9. |
- Gong, Haiyue, et al.
(författare)
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Ag−Polymer nanocomposites for capture, detection, and destruction of bacteria
- 2019
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Ingår i: ACS Applied Nano Materials. - : American Chemical Society (ACS). - 2574-0970. ; 2, s. 1655-1663
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Tidskriftsartikel (refereegranskat)abstract
- Bacterial infection is one of the major problems for human health. To prevent outbreak of bacteria-caused diseases, early diagnosis of bacterial pathogen and effective destruction of pathogenic microorganisms are in urgent need. In this work, we developed a new multifunctional nanocomposite material that can effectively capture and destroy bacteria. Epoxide-modified nanoparticles were synthesized by microemulsion polymerization and precipitation polymerization. The epoxide groups on the particle surface were reacted with polyethylenimine to introduce cationic amine groups. The amine groups on the nanoparticle surface enhanced the colloidal stability of the particles’ suspension and provided multivalent interactions to bind and destroy the bacteria. After further modification with Ag nanoparticles, the final composite nanomaterial was able to not only capture and destroy Gram-negative bacteria but also allow the bacteria’s fingerprint spectra to be obtained through surface-enhanced Raman scattering.The multifunctional nanoparticles developed in this work offer a new approach toward fast capture, detection, and destruction of pathogenic bacteria.
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| 10. |
- Greving, Imke, et al.
(författare)
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Structural Diversity of Native Major Ampullate, Minor Ampullate, Cylindriform, and Flagelliform Silk Proteins in Solution
- 2020
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Ingår i: Biomacromolecules. - : American Chemical Society (ACS). - 1526-4602 .- 1525-7797. ; 21:8, s. 3387-3393
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Tidskriftsartikel (refereegranskat)abstract
- 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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