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1.
Polymers (Basel) ; 16(12)2024 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-38932044

RESUMO

Bio-based solutions for solid timber gluing have always been a very sensitive topic in wood technology. In this work, we optimize the gluing conditions of a starch-tannin formulation, which allows high performance in dry conditions and resistance to water dipping for 3 h, allowing for the D2 classification to be reached according to EN 204. It was observed that the starch-tannin formulations enhanced their performance by increasing the heating temperature, achieving satisfactory results at 140 °C for 13 min. The proportion of polyphenols in the mixture enhances the water resistance but is only tolerated until 20-30%. In particular, the addition of 10% tannin-hexamine enhances the water-resistant properties of starch for both quebracho and chestnut extract. The application of the jet of cold atmospheric plasma allows for good results with more viscous formulations, increasing their penetration in wood. Solid-state 13C-NMR analysis was also performed, and the spectroscopic information suggests establishing a coordination complex between starch and tannin.

2.
Molecules ; 29(8)2024 Apr 09.
Artigo em Inglês | MEDLINE | ID: mdl-38675518

RESUMO

The structural features and optical properties of supramolecular cyanoiron salts containing bis-pyridinium-4-oxime Toxogonin® (TOXO) as an electron acceptor are presented. The properties of the new TOXO-based cyanoiron materials were probed by employing two cyanoiron platforms: hexacyanoferrate(II), [Fe(CN)6]4- (HCF); and nitroprusside, [Fe(CN)5(NO)]2- (NP). Two water-insoluble inter-ionic donor-acceptor phases were characterized: the as-prepared microcrystalline reddish-brown (TOXO)2[Fe(CN)6]·8H2O (1a) with a medium-responsive, hydrochromic character; and the dark violet crystalline (TOXO)2[Fe(CN)6]·3.5H2O (1cr). Complex 1a, upon external stimulation, transforms to the violet anhydrous phase (TOXO)2[Fe(CN)6] (1b), which upon water uptake transforms back to 1a. Using the NP platform resulted in the water-insoluble crystalline salt TOXO[Fe(CN)5(NO)]·2H2O (2). The structures of 1cr and 2, solved by single-crystal X-ray diffraction, along with a comparative spectroscopic (UV-vis-NIR diffuse reflectance, IR, solid-state MAS-NMR, Mössbauer), thermal, powder X-ray diffraction, and microscopic analysis (SEM, TEM) of the isolated materials, provided insight for the supramolecular binding, electron-accepting, and H-bonding capabilities of TOXO in the self-assembly of these functionalized materials.

3.
Photosynth Res ; 2024 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-38662326

RESUMO

It has been thoroughly documented, by using 31P-NMR spectroscopy, that plant thylakoid membranes (TMs), in addition to the bilayer (or lamellar, L) phase, contain at least two isotropic (I) lipid phases and an inverted hexagonal (HII) phase. However, our knowledge concerning the structural and functional roles of the non-bilayer phases is still rudimentary. The objective of the present study is to elucidate the origin of I phases which have been hypothesized to arise, in part, from the fusion of TMs (Garab et al. 2022 Progr Lipid Res 101,163). We take advantage of the selectivity of wheat germ lipase (WGL) in eliminating the I phases of TMs (Dlouhý et al. 2022 Cells 11: 2681), and the tendency of the so-called BBY particles, stacked photosystem II (PSII) enriched membrane pairs of 300-500 nm in diameter, to form large laterally fused sheets (Dunahay et al. 1984 BBA 764: 179). Our 31P-NMR spectroscopy data show that BBY membranes contain L and I phases. Similar to TMs, WGL selectively eliminated the I phases, which at the same time exerted no effect on the molecular organization and functional activity of PSII membranes. As revealed by sucrose-density centrifugation, magnetic linear dichroism spectroscopy and scanning electron microscopy, WGL disassembled the large laterally fused sheets. These data provide direct experimental evidence on the involvement of I phase(s) in the fusion of stacked PSII membrane pairs, and strongly suggest the role of non-bilayer lipids in the self-assembly of the TM system.

4.
Int J Mol Sci ; 24(17)2023 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-37686239

RESUMO

The G4C2 hexanucleotide repeat expansion in the c9orf72 gene is a major genetic cause of familial amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD), with the formation of G-quadruplexes directly linked to the development of these diseases. Cations play a crucial role in the formation and structure of G-quadruplexes. In this study, we investigated the impact of biologically relevant potassium ions on G-quadruplex structures and utilized 15N-labeled ammonium cations as a substitute for K+ ions to gain further insights into cation binding and exchange dynamics. Through nuclear magnetic resonance spectroscopy and molecular dynamics simulations, we demonstrate that the single d(G4C2) repeat, in the presence of 15NH4+ ions, adopts a tetramolecular G-quadruplex with an all-syn quartet at the 5'-end. The movement of 15NH4+ ions through the central channel of the G-quadruplex, as well as to the bulk solution, is governed by the vacant cation binding site, in addition to the all-syn quartet at the 5'-end. Furthermore, the addition of K+ ions to G-quadruplexes folded in the presence of 15NH4+ ions induces stacking of G-quadruplexes via their 5'-end G-quartets, leading to the formation of stable higher-ordered species.


Assuntos
Esclerose Lateral Amiotrófica , Demência Frontotemporal , Degeneração Lobar Frontotemporal , Quadruplex G , Humanos , Esclerose Lateral Amiotrófica/genética , Cátions , Potássio
5.
Biochimie ; 214(Pt A): 73-82, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37573019

RESUMO

The human telomere oligonucleotide, d[TAGGG(TTAGGG)2TTAGG] (TAGGG), can adopt two distinct 2-G-quartet G-quadruplex structures at pH 7.0 and 5.0, referred to as the TD and KDH+ forms, respectively. By using a combination of NMR and computational techniques, we determined high-resolution structures of both forms, which revealed unique loop architectures, base triples, and base pairs that play a crucial role in the pH-driven structural transformation of TAGGG. Our study demonstrated that TAGGG represents a reversible pH-driven switch system where the stability and pH-induced structural transformation of the G-quadruplexes are influenced by the terminal residues and base triples. Gaining insight into the factors that regulate the formation of G-quadruplexes and their pH-sensitive structural equilibrium holds great potential for the rational design of novel DNA based pH-driven switches. These advancements in understanding create exciting opportunities for applications in the field of nanotechnology, specifically in the development of bio-nano-motors.


Assuntos
Quadruplex G , Humanos , DNA/química , Oligonucleotídeos/química , Espectroscopia de Ressonância Magnética , Telômero , Concentração de Íons de Hidrogênio , Conformação de Ácido Nucleico
6.
Molecules ; 28(6)2023 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-36985772

RESUMO

This study examined the potential of hydrolysable tannin in comparison to condensed tannins for the production of furanic foams. The results indicate that chestnut tannin presents lower reactivity and requires a stronger acid for the polymerization. Additionally, foamability and density were found to be dependent on both surfactant concentration and tannin type, allowing lower densities for mimosa tannin and lower thermal conductivities for chestnut-based foams. Mimosa tannin was found to have the highest compression strength, followed by quebracho and chestnut, promising thermal conductivity of around 50 mW/m·K for 300 kg/m3 foams, which suggests that chestnut foams have the potential to performing highly when the density is reduced. Chemical analysis revealed that the methylene moieties of the furanics are non-specific and produces new covalent bonds with nucleophilic substrates: -OH groups and free-positions in the flavonoids. Overall, this study opens new perspectives for the application of hydrolysable tannins in polymer and material science.

7.
Cells ; 11(17)2022 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-36078087

RESUMO

It is well established that plant thylakoid membranes (TMs), in addition to a bilayer, contain two isotropic lipid phases and an inverted hexagonal (HII) phase. To elucidate the origin of non-bilayer lipid phases, we recorded the 31P-NMR spectra of isolated spinach plastoglobuli and TMs and tested their susceptibilities to lipases and proteases; the structural and functional characteristics of TMs were monitored using biophysical techniques and CN-PAGE. Phospholipase-A1 gradually destroyed all 31P-NMR-detectable lipid phases of isolated TMs, but the weak signal of isolated plastoglobuli was not affected. Parallel with the destabilization of their lamellar phase, TMs lost their impermeability; other effects, mainly on Photosystem-II, lagged behind the destruction of the original phases. Wheat-germ lipase selectively eliminated the isotropic phases but exerted little or no effect on the structural and functional parameters of TMs-indicating that the isotropic phases are located outside the protein-rich regions and might be involved in membrane fusion. Trypsin and Proteinase K selectively suppressed the HII phase-suggesting that a large fraction of TM lipids encapsulate stroma-side proteins or polypeptides. We conclude that-in line with the Dynamic Exchange Model-the non-bilayer lipid phases of TMs are found in subdomains separated from but interconnected with the bilayer accommodating the main components of the photosynthetic machinery.


Assuntos
Bicamadas Lipídicas , Tilacoides , Lipase/metabolismo , Bicamadas Lipídicas/metabolismo , Espectroscopia de Ressonância Magnética , Peptídeo Hidrolases/metabolismo , Tilacoides/metabolismo
8.
Nat Commun ; 13(1): 1062, 2022 02 25.
Artigo em Inglês | MEDLINE | ID: mdl-35217667

RESUMO

A possible engineering of materials with diverse bio- and nano-applications relies on robust self-assembly of oligonucleotides. Bottom-up approach utilizing guanine-rich DNA oligonucleotides can lead to formation of G-wires, nanostructures consisting of continuous stacks of G-quartets. However, G-wire structure and self-assembly process remain poorly understood, although they are crucial for optimizing properties needed for specific applications. Herein, we use nuclear magnetic resonance to get insights at molecular level on how chosen short, guanine-rich oligonucleotides self-assemble into G-wires, whereas complementary methods are used for their characterization. Additionally, unravelling mechanistic details enable us to guide G-wire self-assembly in a controlled manner. MD simulations provide insight why loop residues with considerably different properties, i.e., hydrogen-bond affinity, stacking interactions, electronic effects and hydrophobicity extensively increase or decrease G-wire length. Our results provide fundamental understanding of G-wire self-assembly process useful for future design of nanomaterials with specific properties.


Assuntos
Quadruplex G , Nanoestruturas , DNA/química , Guanina , Nanoestruturas/química , Oligonucleotídeos
9.
Polymers (Basel) ; 13(24)2021 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-34960963

RESUMO

One of the major challenges currently in the field of material science is finding natural alternatives to the high-performing plastics developed in the last century. Consumers trust synthetic products for their excellent properties, but they are becoming aware of their impact on the planet. One of the most attractive precursors for natural polymers is tannin extracts and in particular condensed tannins. Quebracho (Schinopsis balansae) extract is one of the few industrially available flavonoids and can be exploited as a building block for thermoset resins due to its phenol-like reactivity. The aim of this study was to systematically investigate different hardeners and evaluate the water resistance, thermal behavior, and chemical structure of the quebracho tannin-based polymers in order to understand their suitability as adhesives. It was observed that around 80% of the extract is resistant to leaching when 5% of formaldehyde or hexamine or 10% of glyoxal or furfural are added. Additionally, furfuryl alcohol guarantees high leaching resistance, but only at higher proportions (20%). The quebracho-based formulations showed specific thermal behavior during hardening and higher degradation resistance than the extract. Finally, these polymers undergo similar chemistry to those of mimosa, with exclusive reactivity of the A-ring of the flavonoid.

10.
Polymers (Basel) ; 13(18)2021 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-34577958

RESUMO

With increasing demand of alternatives to oil-based lightweight materials, the development of tannin-based foams is getting more and more attention. In this paper, an alternative to traditionally used solvent-evaporation in the production of tannin-foams is presented. Mixing the tannin-furanic resin with different amounts of ionic and non-ionic surfactants at high agitational speed allows for the formation of highly porous, mechanically stable tannin-foams. Investigations on the influence of surfactant type and ingredient ratios on the foaming behavior and properties of the final foams were conducted. Materials obtained via this route do present extraordinary compression resistance (about 0.8 MPa), good thermal insulation (40 mW/m·K) and are suitable as a wastewater treatment agent at the end-of-life. It was shown that during mechanical blowing, homogeneous cross-sections and almost perfectly round pores form, leading to the high compression resistance. Investigations by means of Fourier transform infrared and 13C nuclear magnetic resonance spectroscopy show that the milder reaction environment leads to more linear poly(furfuryl alcohol)-tannin chains. This new type of tannin foam allows for use in various different fields of application ranging from durable building insulation to wastewater treatment.

11.
Cells ; 10(9)2021 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-34572003

RESUMO

Build-up of the energized state of thylakoid membranes and the synthesis of ATP are warranted by organizing their bulk lipids into a bilayer. However, the major lipid species of these membranes, monogalactosyldiacylglycerol, is a non-bilayer lipid. It has also been documented that fully functional thylakoid membranes, in addition to the bilayer, contain an inverted hexagonal (HII) phase and two isotropic phases. To shed light on the origin of these non-lamellar phases, we performed 31P-NMR spectroscopy experiments on sub-chloroplast particles of spinach: stacked, granum and unstacked, stroma thylakoid membranes. These membranes exhibited similar lipid polymorphism as the whole thylakoids. Saturation transfer experiments, applying saturating pulses at characteristic frequencies at 5 °C, provided evidence for distinct lipid phases-with component spectra very similar to those derived from mathematical deconvolution of the 31P-NMR spectra. Wheat-germ lipase treatment of samples selectively eliminated the phases exhibiting sharp isotropic peaks, suggesting easier accessibility of these lipids compared to the bilayer and the HII phases. Gradually increasing lipid exchanges were observed between the bilayer and the two isotropic phases upon gradually elevating the temperature from 5 to 35 °C, suggesting close connections between these lipid phases. Data concerning the identity and structural and functional roles of different lipid phases will be presented in the accompanying paper.


Assuntos
Cloroplastos/química , Lipídeos de Membrana/química , Tilacoides/química , Galactolipídeos/química , Espectroscopia de Ressonância Magnética/métodos , Temperatura
12.
J Med Chem ; 64(11): 7809-7838, 2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-34043358

RESUMO

We report on the design, synthesis, and biological evaluation of a series of nucleotide-binding oligomerization-domain-containing protein 2 (NOD2) desmuramylpeptide agonists with improved in vitro and in vivo adjuvant properties. We identified two promising compounds: 68, a potent nanomolar in vitro NOD2 agonist, and the more lipophilic 75, which shows superior adjuvant activity in vivo. Both compounds had immunostimulatory effects on peripheral blood mononuclear cells at the protein and transcriptional levels, and augmented dendritic-cell-mediated activation of T cells, while 75 additionally enhanced the cytotoxic activity of peripheral blood mononuclear cells against malignant cells. The C18 lipophilic tail of 75 is identified as a pivotal structural element that confers in vivo adjuvant activity in conjunction with a liposomal delivery system. Accordingly, liposome-encapsulated 75 showed promising adjuvant activity in mice, surpassing that of muramyl dipeptide, while achieving a more balanced Th1/Th2 immune response, thus highlighting its potential as a vaccine adjuvant.


Assuntos
Acetilmuramil-Alanil-Isoglutamina/química , Adjuvantes Imunológicos/química , Proteína Adaptadora de Sinalização NOD2/agonistas , Acetilmuramil-Alanil-Isoglutamina/metabolismo , Acetilmuramil-Alanil-Isoglutamina/farmacologia , Adjuvantes Imunológicos/metabolismo , Adjuvantes Imunológicos/farmacologia , Animais , Formação de Anticorpos/efeitos dos fármacos , Linhagem Celular , Desenho de Fármacos , Humanos , Imunoglobulina G/metabolismo , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Lipossomos/química , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Proteína Adaptadora de Sinalização NOD2/metabolismo , Ovalbumina/imunologia , Relação Estrutura-Atividade , Células Th1/citologia , Células Th1/imunologia , Células Th1/metabolismo , Células Th2/citologia , Células Th2/imunologia , Células Th2/metabolismo
13.
Sci Rep ; 10(1): 11959, 2020 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-32686730

RESUMO

The role of non-bilayer lipids and non-lamellar lipid phases in biological membranes is an enigmatic problem of membrane biology. Non-bilayer lipids are present in large amounts in all membranes; in energy-converting membranes they constitute about half of their total lipid content-yet their functional state is a bilayer. In vitro experiments revealed that the functioning of the water-soluble violaxanthin de-epoxidase (VDE) enzyme of plant thylakoids requires the presence of a non-bilayer lipid phase. 31P-NMR spectroscopy has provided evidence on lipid polymorphism in functional thylakoid membranes. Here we reveal reversible pH- and temperature-dependent changes of the lipid-phase behaviour, particularly the flexibility of isotropic non-lamellar phases, of isolated spinach thylakoids. These reorganizations are accompanied by changes in the permeability and thermodynamic parameters of the membranes and appear to control the activity of VDE and the photoprotective mechanism of non-photochemical quenching of chlorophyll-a fluorescence. The data demonstrate, for the first time in native membranes, the modulation of the activity of a water-soluble enzyme by a non-bilayer lipid phase.


Assuntos
Bicamadas Lipídicas/química , Oxirredutases/metabolismo , Tilacoides/química , Água/química , Varredura Diferencial de Calorimetria , Compostos de Epóxi/metabolismo , Concentração de Íons de Hidrogênio , Cinética , Luz , Lipídeos/química , Espectroscopia de Ressonância Magnética , Solubilidade , Spinacia oleracea/metabolismo , Temperatura , Xantofilas/metabolismo
14.
Materials (Basel) ; 13(7)2020 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-32244686

RESUMO

The molecular structure of the 8-hydroxyquinoline-bis (2-phenylpyridyl) iridium (IrQ(ppy)2) dual emitter organometallic compound is determined based on detailed 1D and 2D nuclear magnetic resonance (NMR), to identify metal-ligands coordination, isomerization and chemical yield of the desired compound. Meanwhile, the extended X-ray absorption fine structure (EXAFS) was used to determine the interatomic distances around the iridium ion. From the NMR results, this compound IrQ(ppy)2 exhibits a trans isomerization with a distribution of coordinated N-atoms in a similar way to facial Ir(ppy)3. The EXAFS measurements confirm the structural model of the IrQ(ppy)2 compound where the oxygen atoms from the quinoline ligands induce the splitting of the next-nearest neighboring C in the second shell of the Ir3+ ions. The high-performance liquid chromatography (HPLC), as a part of the detailed molecular analysis, confirms the purity of the desired IrQ(ppy)2 organometallic compound as being more than 95%, together with the progress of the chemical reactions towards the final compound. The theoretical model of the IrQ(ppy)2, concerning the expected bond lengths, is compared with the structural model from the EXAFS and XRD measurements.

15.
Molecules ; 25(3)2020 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-31991801

RESUMO

The hexanucleotide expansion GGGGCC located in C9orf72 gene represents the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar dementia (FTLD). Since the discovery one of the non-exclusive mechanisms of expanded hexanucleotide G4C2 repeats involved in ALS and FTLD is RNA toxicity, which involves accumulation of pathological sense and antisense RNA transcripts. Formed RNA foci sequester RNA-binding proteins, causing their mislocalization and, thus, diminishing their biological function. Therefore, structures adopted by pathological RNA transcripts could have a key role in pathogenesis of ALS and FTLD. Utilizing NMR spectroscopy and complementary methods, we examined structures adopted by both guanine-rich sense and cytosine-rich antisense RNA oligonucleotides with four hexanucleotide repeats. While both oligonucleotides tend to form dimers and hairpins, the equilibrium of these structures differs with antisense oligonucleotide being more sensitive to changes in pH and sense oligonucleotide to temperature. In the presence of K+ ions, guanine-rich sense RNA oligonucleotide also adopts secondary structures called G-quadruplexes. Here, we also observed, for the first time, that antisense RNA oligonucleotide forms i-motifs under specific conditions. Moreover, simultaneous presence of sense and antisense RNA oligonucleotides promotes formation of heterodimer. Studied structural diversity of sense and antisense RNA transcripts not only further depicts the complex nature of neurodegenerative diseases but also reveals potential targets for drug design in treatment of ALS and FTLD.


Assuntos
Conformação de Ácido Nucleico , Oligonucleotídeos Antissenso/química , Oligonucleotídeos/química , RNA Antissenso/química , Sequências Repetitivas de Ácido Nucleico , Esclerose Lateral Amiotrófica/etiologia , Pareamento de Bases , Proteína C9orf72/química , Proteína C9orf72/genética , Suscetibilidade a Doenças , Demência Frontotemporal/etiologia , Humanos , Concentração de Íons de Hidrogênio , Espectroscopia de Ressonância Magnética , Oligonucleotídeos/genética , Oligonucleotídeos Antissenso/genética , RNA Antissenso/genética , Análise Espectral , Relação Estrutura-Atividade , Temperatura
16.
Nucleic Acids Res ; 48(4): 2189-2197, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-31950178

RESUMO

Understanding the mechanism by which biological macromolecules fold into their functional native conformations represents a problem of fundamental interest. DNA oligonucleotides derived from human telomeric repeat d[TAGGG(TTAGGG)3] and d[TAGGG(TTAGGG)3TT] fold into G-quadruplexes through diverse steps. Varying the pH and temperature by the use of nuclear magnetic resonance and other methods enabled detection of pre-folded structures that exist in solution before completely formed G-quadruplexes upon addition of cations. Pre-folded structures are in general hard to detect, however their knowledge is crucial to set up folding pathways into final structure since they are believed to be a starting point. Unexpectedly well-defined pre-folded structures composed of base triples for both oligonucleotides were detected at certain pH and temperature. These kinds of structures were up to now only hypothesized as intermediates in the folding process. All revealed pre-folded structures irrespective of the pH and temperature exhibited one common structural feature that could govern folding process.


Assuntos
DNA/química , Quadruplex G , Conformação de Ácido Nucleico , Telômero/genética , Dicroísmo Circular , DNA/genética , Humanos , Concentração de Íons de Hidrogênio , Ligantes , Modelos Moleculares , Oligonucleotídeos/química , Oligonucleotídeos/genética , Telômero/química , Temperatura
17.
Nucleic Acids Res ; 48(5): 2749-2761, 2020 03 18.
Artigo em Inglês | MEDLINE | ID: mdl-31996902

RESUMO

GCn and GCnCG, where n = (G2AG4AG2), fold into well-defined, dimeric G-quadruplexes with unprecedented folding topologies in the presence of Na+ ions as revealed by nuclear magnetic resonance spectroscopy. Both G-quadruplexes exhibit unique combination of structural elements among which are two G-quartets, A(GGGG)A hexad and GCGC-quartet. Detailed structural characterization uncovered the crucial role of 5'-GC ends in formation of GCn and GCnCG G-quadruplexes. Folding in the presence of 15NH4+ and K+ ions leads to 3'-3' stacking of terminal G-quartets of GCn G-quadruplexes, while 3'-GC overhangs in GCnCG prevent dimerization. Results of the present study expand repertoire of possible G-quadruplex structures. This knowledge will be useful in DNA sequence design for nanotechnological applications that may require specific folding topology and multimerization properties.


Assuntos
Composição de Bases/genética , Cátions/metabolismo , DNA/química , Quadruplex G , Dimerização , Modelos Moleculares , Ressonância Magnética Nuclear Biomolecular
18.
Nucleic Acids Res ; 47(21): 11057-11068, 2019 12 02.
Artigo em Inglês | MEDLINE | ID: mdl-31665504

RESUMO

I-motifs are non-canonical nucleic acids structures characterized by intercalated H-bonds between hemi-protonated cytosines. Evidence on the involvement of i-motif structures in the regulation of cellular processes in human cells has been consistently growing in the recent years. However, i-motifs within non-human genomes have never been investigated. Here, we report the characterization of i-motifs within the long terminal repeat (LTR) promoter of the HIV-1 proviral genome. Biophysical and biochemical analysis revealed formation of a predominant i-motif with an unprecedented loop composition. One-dimensional nuclear magnetic resonance investigation demonstrated formation of three G-C H-bonds in the long loop, which likely improve the structure overall stability. Pull-down experiments combined with mass spectrometry and protein crosslinking analysis showed that the LTR i-motif is recognized by the cellular protein hnRNP K, which induced folding at physiological conditions. In addition, hnRNP K silencing resulted in an increased LTR promoter activity, confirming the ability of the protein to stabilize the i-motif-forming sequence, which in turn regulates the LTR-mediated HIV-1 transcription. These findings provide new insights into the complexity of the HIV-1 virus and lay the basis for innovative antiviral drug design, based on the possibility to selectively recognize and target the HIV-1 LTR i-motif.


Assuntos
Repetição Terminal Longa de HIV , HIV-1 , Ribonucleoproteínas Nucleares Heterogêneas Grupo K/química , Regiões Promotoras Genéticas , Provírus , RNA Viral/química , Sítios de Ligação , Regulação Viral da Expressão Gênica , HIV-1/genética , HIV-1/fisiologia , Provírus/genética , Provírus/fisiologia , Transcrição Gênica , Replicação Viral
19.
Acta Pharm ; 69(1): 17-32, 2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31259719

RESUMO

Ibuprofen, a weakly acidic non-steroidal anti-inflammatory drug having poor aqueous solubility, is a challenging drug for the development of pharmaceutical formulations, resulting in numerous research attempts focusing on improvement of its solubility and consequently bioavailability. Most studies have been done for solid dosage forms, with very little attention paid to parenterals. Hence, the main purpose of the present study was to enhance ibuprofen solubility as a result of formulation composition and the freeze drying process. Moreover, the purpose was to prepare a freeze dried dosage form with improved ibuprofen solubility that could, after simple reconstitution with water for injection, result in an isotonic parenteral solution. Solubility of ibuprofen was modified by various excipients suitable for parenteral application. Drug interactions with selected excipients in the final product/lyophilisate were studied by a combined use of XRPD, DSC, Raman and ss-NMR. Analyses of lyophilized samples showed solubility enhancement of ibuprofen and in situ formation of an ibuprofen salt with the alkaline excipients used.


Assuntos
Ibuprofeno/química , Solubilidade/efeitos dos fármacos , Anti-Inflamatórios não Esteroides/química , Disponibilidade Biológica , Varredura Diferencial de Calorimetria/métodos , Química Farmacêutica/métodos , Cristalização/métodos , Formas de Dosagem , Excipientes/química , Liofilização/métodos , Pós/química , Água/química , Difração de Raios X/métodos
20.
Physiol Plant ; 166(1): 278-287, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30666653

RESUMO

Earlier experiments, using 31 P-NMR and time-resolved merocyanine fluorescence spectroscopy, have shown that isolated intact, fully functional plant thylakoid membranes, in addition to the bilayer phase, contain three non-bilayer (or non-lamellar) lipid phases. It has also been shown that the lipid polymorphism of thylakoid membranes can be characterized by remarkable plasticity, i.e. by significant variations in 31 P-NMR signatures. However, changes in the lipid-phase behaviour of thylakoids could not be assigned to changes in the overall membrane organization and the photosynthetic activity, as tested by circular dichroism and 77 K fluorescence emission spectroscopy and the magnitude of the variable fluorescence of photosystem II, which all showed only marginal variations. In this work, we investigated in more detail the temporal stability of the different lipid phases by recording 31 P-NMR spectra on isolated thylakoid membranes that were suspended in sorbitol- or NaCl-based media. We observed, at 5°C during 8 h in the dark, substantial gradual enhancement of the isotropic lipid phases and diminishment of the bilayer phase in the sorbitol-based medium. These changes compared well with the gradually increasing membrane permeability, as testified by the gradual acceleration of the decay of flash-induced electrochromic absorption changes and characteristic changes in the kinetics of fast chlorophyll a-fluorescence transients; all variations were much less pronounced in the NaCl-based medium. These observations suggest that non-bilayer lipids and non-lamellar lipid phases play significant roles in the structural dynamics and functional plasticity of thylakoid membranes.


Assuntos
Membranas Intracelulares/metabolismo , Bicamadas Lipídicas/metabolismo , Tilacoides/metabolismo , Cinética , Espectroscopia de Ressonância Magnética , Complexo de Proteína do Fotossistema II/metabolismo
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