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1.
J Mammary Gland Biol Neoplasia ; 29(1): 10, 2024 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-38722417

RESUMO

Signal transducers and activators of transcription (STAT) proteins regulate mammary development. Here we investigate the expression of phosphorylated STAT3 (pSTAT3) in the mouse and cow around the day of birth. We present localised colocation analysis, applicable to other mammary studies requiring identification of spatially congregated events. We demonstrate that pSTAT3-positive events are multifocally clustered in a non-random and statistically significant fashion. Arginase-1 expressing cells, consistent with macrophages, exhibit distinct clustering within the periparturient mammary gland. These findings represent a new facet of mammary STAT3 biology, and point to the presence of mammary sub-microenvironments.


Assuntos
Células Epiteliais , Glândulas Mamárias Animais , Fator de Transcrição STAT3 , Animais , Feminino , Bovinos , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Animais/citologia , Glândulas Mamárias Animais/crescimento & desenvolvimento , Camundongos , Células Epiteliais/metabolismo , Fator de Transcrição STAT3/metabolismo , Fosforilação , Gravidez , Parto/fisiologia , Parto/metabolismo , Transdução de Sinais
2.
Langmuir ; 35(25): 8344-8356, 2019 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-31122018

RESUMO

For evolving biological and biomedical applications of hybrid protein?lipid materials, understanding the behavior of the protein within the lipid mesophase is crucial. After more than two decades since the invention of the in meso crystallization method, a protein-eye view of its mechanism is still lacking. Numerous structural studies have suggested that integral membrane proteins preferentially partition at localized flat points on the bilayer surface of the cubic phase with crystal growth occurring from a local fluid lamellar L? phase conduit. However, studies to date have, by necessity, focused on structural transitions occurring in the lipid mesophase. Here, we demonstrate using small-angle neutron scattering that the lipid bilayer of monoolein (the most commonly used lipid for in meso crystallization) can be contrast-matched using deuteration, allowing us to isolate scattering from encapsulated peptides during the crystal growth process for the first time. During in meso crystallization, a clear decrease in form factor scattering intensity of the peptides was observed and directly correlated with crystal growth. A transient fluid lamellar L? phase was observed, providing direct evidence for the proposed mechanism for this technique. This suggests that the peptide passes through a transition from the cubic QII phase, via an L? phase to the lamellar crystalline Lc phase with similar layered spacing. When high protein loading was possible, the lamellar crystalline Lc phase of the peptide in the single crystals was observed. These findings show the mechanism of in meso crystallization for the first time from the perspective of integral membrane proteins.


Assuntos
Cristalização/métodos , Bicamadas Lipídicas/química , Glicerídeos/química , Difração de Raios X
3.
Phys Chem Chem Phys ; 21(46): 25649-25657, 2019 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-31723955

RESUMO

The effects of adding silica nanoparticles of varying size and surface chemistry to a liquid crystal system were analysed using small-angle scattering and polarising light microscopy, with varying temperature and applied shear. It was found that nanoparticles aggregate at domain boundaries, causing a reduction in average liquid crystal domain size. These particles can inhibit phase transitions that occur at specific temperatures, ascribed to aggregates posing a kinetic barrier to rearrangement required for phase transitions. Nanoparticles can also promote the existence of specific phases, such as a deswollen hexagonal mesophase for the system studied here, suggested to be caused by silica aggregates 'templating' new phases. These findings have important implications for the application of such systems in biotechnology, and particularly the ability to completely inhibit a phase change at low temperature suggests the potential for mechanistic insight into new methods of cryopreservation.

4.
Langmuir ; 34(31): 9238-9251, 2018 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-29989819

RESUMO

The development of protein-based hydrogels for tissue engineering applications is often limited by their mechanical properties. Herein, we present the facile fabrication of tough regenerated silk fibroin (RSF)/graphene oxide (GO) nanocomposite hydrogels by a photochemical cross-linking method. The RSF/GO composite hydrogels demonstrated soft and adhesive properties during initial stages of photocrosslinking (<2 min), which is not observed for the pristine RSF hydrogel, and rendered a tough and nonadhesive hydrogel upon complete cross-linking (10 min). The composite hydrogels exhibited superior tensile mechanical properties, increased ß-sheet content, and decreased chain mobility compared to that of the pristine RSF hydrogels. The composite hydrogels demonstrated Young's modulus as high as ∼8 MPa, which is significantly higher than native cartilage (∼1.5 MPa), and tensile toughness as high as ∼2.4 MJ/m3, which is greater than that of electroactive polymer muscles and at par with RSF/GO composite membranes fabricated by layer-by-layer assembly. Small-angle scattering study reveals the hierarchical structure of photocrosslinked RSF hydrogels to comprise randomly distributed water-poor (hydrophobic) and water-rich (hydrophilic) regions at the nanoscale, whereas water pores and channels exhibiting fractal-like characteristics at the microscale. The size of hydrophobic domain (containing ß-sheets) was observed to increase slightly with GO incorporation and/or alcohol post-treatment, whereas the size of the hydrophilic domain (intersheet distance containing random coils) was observed to increase significantly, which influences/affects water uptake capacity, cross-link density, and mechanical properties of hydrogels. The presented results have implications for both fundamental understanding of the structure-property relationship of RSF-based hydrogels and their technological applications.

5.
Proteins ; 85(7): 1371-1378, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28380660

RESUMO

The anti-cancer complex, Bovine Alpha-lactalbumin Made LEthal to Tumors (BAMLET), has intriguing broad-spectrum anti-cancer activity. Although aspects of BAMLET's anti-cancer mechanism are still not known, it is understood that it involves the oleic acid or oleate component of BAMLET being preferentially released into cancer cell membranes leading to increased membrane permeability and lysis. The structure of the protein component of BAMLET has previously been elucidated by small angle X-ray scattering (SAXS) to be partially unfolded and dramatically enlarged. However, the structure of the oleic acid component of BAMLET and its disposition with respect to the protein component was not revealed as oleic acid has the same X-ray scattering length density (SLD) as water. Employing the difference in the neutron SLDs of hydrogen and deuterium, we carried out solvent contrast variation small angle neutron scattering (SANS) experiments of hydrogenated BAMLET in deuterated water buffers, to reveal the size, shape, and disposition of the oleic acid component of BAMLET. Our resulting analysis and models generated from SANS and SAXS data indicate that oleic acid forms a spherical droplet of oil incompletely encapsulated by the partially unfolded protein component. This model provides insight into the anti-cancer mechanism of this cache of lipid. The model also reveals a protein component "tail" not associated with the oleic acid component that is able to interact with the tail of other BAMLET molecules, providing a plausible explanation of how BAMLET readily forms aggregates. Proteins 2017; 85:1371-1378. © 2017 Wiley Periodicals, Inc.


Assuntos
Antineoplásicos/química , Deutério/química , Hidrogênio/química , Lactalbumina/química , Ácido Oleico/química , Humanos , Hidrogenação , Conformação Molecular , Difração de Nêutrons , Desdobramento de Proteína , Espalhamento a Baixo Ângulo , Difração de Raios X
6.
Biophys J ; 110(1): 85-94, 2016 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-26745412

RESUMO

The formation of amyloid deposits is a common feature of a broad range of diseases, including atherosclerosis, Alzheimer's disease, and Parkinson's disease. The basis and role of amyloid deposition in the pathogenesis of these diseases is still being defined, however an interesting feature of amyloidogenic proteins is that the majority of the pathologically associated proteins are involved in lipid homeostasis, be it in lipid transport, incorporation into membranes, or the regulation of lipid pathways. Thus, amyloid-forming proteins commonly bind lipids, and lipids are generally involved in the proper folding of these proteins. However, understanding of the basis for these lipid-related aspects of amyloidogenesis is lacking. Thus, we have used the apolipoprotein C-II amyloid model system in conjunction with x-ray and neutron scattering analyses to address this problem. Apolipoprotein C-II is a well-studied model system of systemic amyloid fibril formation, with a clear and well-defined pathway for fibril formation, where the effects of lipid interaction are characterized, particularly for the lipid mimetic dodecylphosphocholine. We show that the micellar state of an inhibitory lipid can have a very significant effect on protein conformation, with micelles stabilizing a particular α-helical structure, whereas submicellar lipids stabilize a very different dimeric, α-helical structure. These results indicate that lipids may have an important role in the development and progression of amyloid-related diseases.


Assuntos
Amiloide/química , Apolipoproteína C-II/química , Materiais Biomiméticos/farmacologia , Lipídeos/química , Micelas , Fosforilcolina/análogos & derivados , Apolipoproteína C-II/metabolismo , Materiais Biomiméticos/química , Materiais Biomiméticos/metabolismo , Modelos Moleculares , Fosforilcolina/química , Fosforilcolina/metabolismo , Fosforilcolina/farmacologia , Agregados Proteicos/efeitos dos fármacos , Conformação Proteica , Estabilidade Proteica
7.
Mol Pharm ; 12(7): 2305-17, 2015 Jul 06.
Artigo em Inglês | MEDLINE | ID: mdl-25936529

RESUMO

Esophageal cancer patients are often diagnosed as "advanced" cases. These patients are subjected to palliative stenting using self-expanding metallic stents (SEMS) to maintain oral alimentation. Unfortunately, SEMS get reoccluded due to tumor growth, in and over the stent struts. To investigate potential solutions to this problem, docetaxel (DTX) delivery films were prepared using PurSil AL 20 (PUS), which can be used as a covering material for the SEMS. Drug-polymer miscibility and interactions were studied. Bilayer films were prepared by adhering the blank film to the DTX loaded film in order to maintain the unidirectional delivery to the esophagus. In vitro release and the local DTX delivery were studied using in vitro permeation experiments. It was found that DTX and PUS were physically and chemically compatible. The bilayer films exhibited sustained release (>30 days) and minimal DTX permeation through esophageal tissues in vitro. The rate-determining step for the DTX delivery was calculated. It was found that >0.9 fraction of rate control lies with the esophageal tissues, suggesting that DTX delivery can be sustained for longer periods compared to the in vitro release observed. Thus, the bilayer films can be developed as a localized sustained delivery system in combination with the stent.


Assuntos
Neoplasias Esofágicas/tratamento farmacológico , Esôfago/efeitos dos fármacos , Taxoides/administração & dosagem , Taxoides/química , Animais , Preparações de Ação Retardada/química , Preparações de Ação Retardada/farmacologia , Docetaxel , Sistemas de Liberação de Medicamentos/métodos , Polímeros/química , Stents , Suínos
8.
Langmuir ; 31(32): 8882-91, 2015 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-26177160

RESUMO

The ability to tune the thermoresponsiveness of recombinant resilin protein, Rec1-resilin, through a facile coassembly system was investigated in this study. The effects of change in conformation and morphology with time and the responsive behavior of Rec1-resilin in solution were studied in response to the addition of a rigid model polypeptide (poly-l-proline) or a hydrophobic rigid protein (Bombyx mori silk fibroin). It was observed that by inducing more ordered conformations and increasing the hydrophobicity the lower critical solution temperature (LCST) of the system was tuned to lower values. Time and temperature were found to be critical parameters in controlling the coassembly behavior of Rec1-resilin in both the model polypeptide and more complex protein systems. Such unique properties are useful for a wide range of applications, including drug delivery and soft tissue engineering applications.


Assuntos
Fibroínas/química , Proteínas de Insetos/química , Peptídeos/química , Temperatura , Animais , Bombyx , Interações Hidrofóbicas e Hidrofílicas
9.
J Pharm Pharm Sci ; 18(4): 773-824, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26626257

RESUMO

The HAMLET family of compounds (Human Alpha-lactalbumin Made Lethal to Tumours) was discovered during studies on the properties of human milk, and is a class of protein-lipid complexes having broad spectrum anti-cancer, and some specific anti-bacterial properties. The structure of HAMLET-like compounds consists of an aggregation of partially unfolded protein making up the majority of the compound's mass, with fatty acid molecules bound in the hydrophobic core. This is a novel protein-lipid structure and has only recently been derived by small-angle X-ray scattering analysis. The structure is the basis of a novel cytotoxicity mechanism responsible for anti-cancer activity to all of the around 50 different cancer cell types for which the HAMLET family has been trialled. Multiple cytotoxic mechanisms have been hypothesised for the HAMLET-like compounds, but it is not yet clear which of those are the initiating cytotoxic mechanism(s) and which are subsequent activities triggered by the initiating mechanism(s). In addition to the studies into the structure of these compounds, this review presents the state of knowledge of the anti-cancer aspects of HAMLET-like compounds, the HAMLET-induced cytotoxic activities to cancer and non-cancer cells, and the several prospective cell membrane and intracellular targets of the HAMLET family. The emerging picture is that HAMLET-like compounds initiate their cytotoxic effects on what may be a cancer-specific target in the cell membrane that has yet to be identified. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.


Assuntos
Lactalbumina/farmacologia , Leite Humano/química , Neoplasias/tratamento farmacológico , Ácidos Oleicos/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Humanos , Interações Hidrofóbicas e Hidrofílicas , Lactalbumina/química , Lactalbumina/isolamento & purificação , Neoplasias/patologia , Ácidos Oleicos/química , Ácidos Oleicos/isolamento & purificação
10.
Proteins ; 82(7): 1400-8, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24408789

RESUMO

BAMLET (Bovine Alpha-lactalbumin Made LEthal to Tumors) is a member of the family of the HAMLET-like complexes, a novel class of protein-based anti-cancer complexes that incorporate oleic acid and deliver it to cancer cells. Small angle X-ray scattering (SAXS) was performed on the complex at pH 12, examining the high pH structure as a function of oleic acid added. The SAXS data for BAMLET species prepared with a range of oleic acid concentrations indicate extended, irregular, partially unfolded protein conformations that vary with the oleic acid concentration. Increases in oleic acid concentration correlate with increasing radius of gyration without an increase in maximum particle dimension, indicating decreasing protein density. The models for the highest oleic acid content BAMLET indicate an unusual coiled elongated structure that contrasts with apo-α-lactalbumin at pH 12, which is an elongated globular molecule, suggesting that oleic acid inhibits the folding or collapse of the protein component of BAMLET to the globular form. Circular dichroism of BAMLET and apo-α-lactalbumin was performed and the results suggest that α-lactalbumin and BAMLET unfold in a continuum of increasing degree of unfolded states. Taken together, these results support a model in which BAMLET retains oleic acid by non-specific association in the core of partially unfolded protein, and represent a new type of lipoprotein structure.


Assuntos
Lactalbumina/química , Ácido Oleico/química , Animais , Bovinos , Dicroísmo Circular , Concentração de Íons de Hidrogênio , Modelos Moleculares , Espalhamento a Baixo Ângulo , Difração de Raios X
11.
Proteins ; 82(1): 10-21, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23733394

RESUMO

Alpha-synuclein (α-syn) forms the amyloid-containing Lewy bodies found in the brain in Parkinson's disease. The neurotransmitter dopamine (DA) reacts with α-syn to form SDS-resistant soluble, non-amyloid, and melanin-containing oligomers. Their toxicity is debated, as is the nature of their structure and their relation to amyloid-forming conformers of α-syn. The small-angle X-ray scattering technique in combination with modeling by the ensemble optimization method showed that the un-reacted native protein populated three broad classes of conformer, while reaction with DA gave a restricted ensemble range suggesting that the rigid melanin molecule played an important part in their structure. We found that 6 M guanidine hydrochloride did not dissociate α-syn DA-reacted dimers and trimers, suggesting covalent linkages. The pathological significance of covalent association is that if they are non-toxic, the oligomers would act as a sink for toxic excess DA and α-syn; if toxic, their stability could enhance their toxicity. We argue it is essential, therefore, to resolve the question of whether they are toxic or not.


Assuntos
Encéfalo/metabolismo , Dopamina/metabolismo , Guanidina/metabolismo , Doença de Parkinson/metabolismo , Desnaturação Proteica , alfa-Sinucleína/metabolismo , Análise por Conglomerados , Eletroforese em Gel de Poliacrilamida , Escherichia coli , Humanos , Modelos Químicos , Espalhamento de Radiação , Ultracentrifugação
12.
AoB Plants ; 16(2): plae023, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38638333

RESUMO

Recent changes in water availability can be crucial for the development, growth and carbon budget of forests. Therefore, our aim was to determine the effect of reduced throughfall and severe summer drought on stem CO2 efflux as a function of temperature and stem increment. Stem CO2 efflux was measured using the chamber method on oak and hornbeam under four treatments: coppice, thinned coppice, and both coppice and thinned coppice with 30 %-reduced throughfall. The first year of the experiment had favourable soil water availability and the second year was characterized by a dry summer. While reduced throughfall had no effect on stem CO2 efflux, the summer drought decreased efflux by 43-81 % during July and August. The stem CO2 efflux was reduced less severely (by 13-40 %) in September when the drought persisted but the stem increment was already negligible. The stem increment was also strongly affected by the drought, which was reflected in its paired relationship with stem CO2 efflux over the two experimental years. The study showed that summer dry periods significantly and rapidly reduce stem CO2 efflux, whereas a constant 30 % rainfall reduction needs probably a longer time to affect stem properties, and indirectly stem CO2 efflux.

13.
J Synchrotron Radiat ; 20(Pt 4): 626-34, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23765306

RESUMO

This paper reports the results of time-resolved synchrotron small-angle scattering and powder diffraction experiments where natrojarosites were synthesized in situ in order to observe the species produced at the earliest stages of nucleation. The sample temperatures were 333, 353 and 368 K. These compounds were synthesized by co-precipitation from solution on the Small and Wide Angle Scattering and Powder Diffraction beamlines at the Australian Synchrotron. Scattering data were collected continuously throughout the syntheses. The results presented here show that the first particles to form in solution appear to be amorphous and nucleate on the walls of the reaction vessel. Crucially, there is a single nucleation event which forms particles with an elliptical disc morphology which then grow uniformly before natrojarosite crystallization is observed in complementary powder diffraction data. This nucleation event may represent the key to controlling the growth of jarosites in industrial and environmental settings.

14.
Biomacromolecules ; 14(11): 3850-60, 2013 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-24047190

RESUMO

In this work, we have applied an electrospinning method to control wettability and further hydrophobic modification of a hydrophobic polymer mat of poly(vinylidene fluoride-co-hexafluoropropylene). A correlation between the processing parameters, rheological properties of polymer solutions, and electrospinning ability was made using the polymer's critical entanglement concentration, the boundary between the semidilute unentangled regime and the semidilute entangled regime. The wetting behavior, structural and thermal characteristics of electrospun (ES) mats were evaluated and compared with solvent cast sample using advancing and receding contact angle analyses, differential scanning calorimetry, and small-angle X-ray scattering. To demonstrate the feasibility, the best optimized ES samples were examined for their potential and ability to support bone marrow derived endothelial cell seeding efficiency, adhesion and proliferation. Our studies show that, while different processing techniques can effectively modulate physical and morphological changes such as porosity and hydrophobicity, the cellular adhesion and proliferation are highly time-dependent and controlled by chemical factors. As such, these results suggest that it is the interplay of both physical and chemical factors that determine the endothelialization of porous near superhydrophobic scaffolds. The developed electrospun samples demonstrate their feasibility for endothelialization.


Assuntos
Células da Medula Óssea/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Interações Hidrofóbicas e Hidrofílicas , Polivinil/farmacologia , Adesão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Células Endoteliais/citologia , Humanos , Estrutura Molecular , Tamanho da Partícula , Polivinil/síntese química , Polivinil/química , Porosidade , Relação Estrutura-Atividade , Propriedades de Superfície , Temperatura , Fatores de Tempo , Molhabilidade
15.
Commun Biol ; 6(1): 124, 2023 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-36721030

RESUMO

Group A Streptococcus (GAS) is a strict human pathogen possessing a unique pathogenic trait that utilizes the cooperative activity of NAD+-glycohydrolase (NADase) and Streptolysin O (SLO) to enhance its virulence. How NADase interacts with SLO to synergistically promote GAS cytotoxicity and intracellular survival is a long-standing question. Here, the structure and dynamic nature of the NADase/SLO complex are elucidated by X-ray crystallography and small-angle scattering, illustrating atomic details of the complex interface and functionally relevant conformations. Structure-guided studies reveal a salt-bridge interaction between NADase and SLO is important to cytotoxicity and resistance to phagocytic killing during GAS infection. Furthermore, the biological significance of the NADase/SLO complex in GAS virulence is demonstrated in a murine infection model. Overall, this work delivers the structure-functional relationship of the NADase/SLO complex and pinpoints the key interacting residues that are central to the coordinated actions of NADase and SLO in the pathogenesis of GAS infection.


Assuntos
Streptococcus , Estreptolisinas , Humanos , Animais , Camundongos , Proteínas de Bactérias , NAD+ Nucleosidase
16.
AIDS Patient Care STDS ; 36(7): 263-271, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35727648

RESUMO

Current expert recommendations suggest anal cytology followed by high-resolution anoscopy (HRA) for biopsy and histological confirmation may be beneficial in cancer prevention, especially in people living with HIV (PLWH). Guided by the social ecological model, the purpose of this study was to examine sociodemographic and clinical variables, individual-level factors (depression, HIV/AIDS-related stigma, and health beliefs) and interpersonal-level factors (social support) related to time to HRA follow-up after abnormal anal cytology. We enrolled 150 PLWH from a large HIV community clinic, with on-site HRA availability, in Atlanta, GA. The median age was 46 years (interquartile range of 37-52), 78.5% identified as African American/Black, and 88.6% identified as born male. The average length of follow-up to HRA after abnormal anal cytology was 380.6 days (standard deviation = 317.23). Only 24.3% (n = 39) of the sample had an HRA within 6 months after an abnormal anal cytology, whereas 57% of the sample had an HRA within 12 months. HIV/AIDS-related stigma [odds ratio (OR) 0.54, 95% confidence interval (CI) 0.33-0.90] and health motivation (OR 0.80, 95% CI 0.67-0.95) were associated with time to HRA follow-up ≤6 months. For HRA follow-up ≤12 months, we found anal cytology [high-grade squamous intraepithelial lesions/atypical squamous cells of undetermined significance cannot exclude HSIL (HSIL/ASCUS-H) vs. low-grade squamous intraepithelial lesions (LSIL) OR = 0.05, 95% CI 0.00-0.70; atypical squamous cells of undetermined significance (ASCUS) vs. LSIL OR = 0.12, 95% CI 0.02-0.64] and health motivation (OR = 0.86, 95% CI 0.65-0.99) were associated. Findings from this study can inform strategies to improve follow-up care after abnormal anal cytology at an individual and interpersonal level in efforts to decrease anal cancer morbidity and mortality.


Assuntos
Neoplasias do Ânus , Células Escamosas Atípicas do Colo do Útero , Infecções por HIV , Infecções por Papillomavirus , Canal Anal/patologia , Neoplasias do Ânus/patologia , Células Escamosas Atípicas do Colo do Útero/patologia , Feminino , Seguimentos , Infecções por HIV/complicações , Homossexualidade Masculina , Humanos , Masculino , Pessoa de Meia-Idade , Infecções por Papillomavirus/complicações
17.
Langmuir ; 26(2): 1239-46, 2010 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-19722605

RESUMO

The immobilization of biological species such as proteins and enzymes in sol-gel hosts is currently an area of intense research activity. However, the majority of these studies have been directed toward investigating the biological activity or physicochemical properties of the encapsulated species, with much less attention having been directed toward the effect of proteins on the structural evolution of the sol-gel matrix. This study investigates the structural evolution of sol-gel matrices in the presence of a model protein, bovine serum albumin (BSA). The sol-gel matrices were produced via the NaF-catalyzed hydrolysis of a mixture of tetramethyoxysilane (TMOS) and methyltrimethoxysilane (MTMS), yielding nanohybrid matrices with controlled pore sizes, pore volumes, and surface chemistry. The structural evolution of the matrix was investigated using a complementary suite of techniques, including solid-state (29)Si NMR, FTIR, SANS contrast variation, and N(2) sorption. A novel approach was developed to model the SANS data, to extract key structural parameters. The results indicated that the structural evolution of the matrices was modulated by a series of complex interactions between the enzyme and the evolving sol-gel nanohybrid: On the molecular scale, increasing BSA content led to an associated increase in both the abundance of linear Si-O-Si species (FTIR) and the Qn network connectivity ((29)Si NMR). However, only minor changes in the connectivity of the evolving Tn network were evident with varying BSA content. The selective role of the protein in these systems, where the approach of the methylated monomer to the vicinity of the protein's surface is presumably impeded by the hydrophobicity of the monomer, will be discussed. On the nanoscale, N(2) sorption data were consistent with an initial increase in the mesopore volume and surface area at low BSA loadings, followed by a subsequent monotonic decrease with increasing BSA content. In contrast, no such trends were evident in the in situ SANS data obtained from these samples, suggesting that modulation of the evolving network structure of the silica matrix by BSA during condensation prevents collapse of the nanoscale gel structure during freeze-drying. This latter comparison reflects the important role of in situ techniques such as small angle scattering (which can be used to study both open and closed porosity and probe nanostructure on length scales from approximately 1 nm to >100 nm) in investigating such complex, multicomponent systems, and techniques for modeling such data in sol-gel systems will be discussed.


Assuntos
Soroalbumina Bovina/química , Animais , Bovinos , Espectroscopia de Ressonância Magnética , Conformação Proteica , Silanos/química , Dióxido de Silício/química , Fluoreto de Sódio/química , Espectroscopia de Infravermelho com Transformada de Fourier
18.
Langmuir ; 26(24): 19073-83, 2010 Dec 21.
Artigo em Inglês | MEDLINE | ID: mdl-21090663

RESUMO

The high-temperature morphology of supported liquid membranes (SLMs) prepared from perfluorinated membranes such as Nafion and Hyflon and hydrophobic ionic liquid 1-butyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide (BMI-TFSI) has been investigated by small-angle X-ray scattering (SAXS). Proton conductivity results of SLMs before and after leaching show an increase in conductivity with temperature up to 160 °C in an anhydrous environment. DSC results show that crystallites within perfluorinated membranes are thermally stable up to 196 °C. High-temperature SAXS results have been used to correlate structure and morphology of supported liquid membranes with high-temperature conductivity data. The ionic liquid essentially acts as a proton solvent in a similar way to water in hydrated Nafion membranes and increases size of clusters, which allow percolation to be achieved more easily. The cation of the ionic liquid interacts with sulfonate groups within ionic domains through electrostatic interactions and displaces protons. Protons can associate with free anions of the ionic liquid, which are loosely associated with cations and can transport by hopping from anion sites within the membrane. The ionic liquid contributes to proton conductivity at high temperature through achievement of long-range ordering and subsequent percolation.

19.
Eur Biophys J ; 39(10): 1407-19, 2010 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-20309679

RESUMO

Inclusions of aggregated alpha-synuclein (alpha-syn) in dopaminergic neurons are a characteristic histological marker of Parkinson's disease (PD). In vitro, alpha-syn in the presence of dopamine (DA) at physiological pH forms SDS-resistant non-amyloidogenic oligomers. We used a combination of biophysical techniques, including sedimentation velocity analysis, small angle X-ray scattering (SAXS) and circular dichroism spectroscopy to study the characteristics of alpha-syn oligomers formed in the presence of DA. Our SAXS data show that the trimers formed by the action of DA on alpha-syn consist of overlapping worm-like monomers, with no end-to-end associations. This lack of structure contrasts with the well-established, extensive beta-sheet structure of the amyloid fibril form of the protein and its pre-fibrillar oligomers. We propose on the basis of these and earlier data that oxidation of the four methionine residues at the C- and N-terminal ends of alpha-syn molecules prevents their end-to-end association and stabilises oligomers formed by cross linking with DA-quinone/DA-melanin, which are formed as a result of the redox process, thus inhibiting formation of the beta-sheet structure found in other pre-fibrillar forms of alpha-syn.


Assuntos
Dopamina/farmacologia , Multimerização Proteica/efeitos dos fármacos , alfa-Sinucleína/química , alfa-Sinucleína/metabolismo , Amiloide/química , Amiloide/metabolismo , Benzoquinonas/química , Benzoquinonas/metabolismo , Dicroísmo Circular , Reagentes de Ligações Cruzadas/farmacologia , Espectroscopia de Ressonância de Spin Eletrônica , Concentração de Íons de Hidrogênio , Melaninas/química , Melaninas/metabolismo , Metionina/química , Metionina/metabolismo , Oxirredução , Doença de Parkinson/metabolismo , Dobramento de Proteína/efeitos dos fármacos , Multimerização Proteica/fisiologia , Estrutura Secundária de Proteína/efeitos dos fármacos , Espalhamento a Baixo Ângulo , Ultracentrifugação
20.
Biosensors (Basel) ; 9(4)2019 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-31671878

RESUMO

Herein we report the first example of a facile biomineralization process to produce ultra-small-sized highly fluorescent aqueous dispersions of platinum noble metal quantum clusters (Pt-NMQCs) using a multi-stimulus responsive, biomimetic intrinsically disordered protein (IDP), Rec1-resilin. We demonstrate that Rec1-resilin acts concurrently as the host, reducing agent, and stabilizer of the blue-green fluorescent Pt-NMQCs once they are being formed. The photophysical properties, quantum yield, and fluorescence lifetime measurements of the synthesized Pt-NMQCs were examined using UV-Vis and fluorescence spectroscopy. The oxidation state of the Pt-NMQCs was quantitatively analyzed using X-ray photoelectron spectroscopy. Both a small angle X-ray scattering technique and a modeling approach have been attempted to present a detailed understanding of the structure and conformational dynamics of Rec1-resilin as an IDP during the formation of the Pt-NMQCs. It has been demonstrated that the green fluorescent Pt-NMQCs exhibit a high quantum yield of ~7.0% and a lifetime of ~9.5 ns in aqueous media. The change in photoluminescence properties due to the inter-dot interactions between proximal dots and aggregation of the Pt-NMQCs by evaporation was also measured spectroscopically and discussed.


Assuntos
Técnicas Biossensoriais , Corantes Fluorescentes/síntese química , Nanopartículas Metálicas/química , Platina/química , Corantes Fluorescentes/química , Espectrometria de Fluorescência
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