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1.
Materials (Basel) ; 17(11)2024 Jun 06.
Artículo en Inglés | MEDLINE | ID: mdl-38894049

RESUMEN

The intensive development of hydrogen technologies has made very promising applications of one of the cheapest and easily produced bulk MgB2-based superconductors. These materials are capable of operating effectively at liquid hydrogen temperatures (around 20 K) and are used as elements in various devices, such as magnets, magnetic bearings, fault current limiters, electrical motors, and generators. These applications require mechanically and chemically stable materials with high superconducting characteristics. This review considers the results of superconducting and structural property studies of MgB2-based bulk materials prepared under different pressure-temperature conditions using different promising methods: hot pressing (30 MPa), spark plasma sintering (16-96 MPa), and high quasi-hydrostatic pressures (2 GPa). Much attention has been paid to the study of the correlation between the manufacturing pressure-temperature conditions and superconducting characteristics. The influence of the amount and distribution of oxygen impurity and an excess of boron on superconducting characteristics is analyzed. The dependence of superconducting characteristics on the various additions and changes in material structure caused by these additions are discussed. It is shown that different production conditions and additions improve the superconducting MgB2 bulk properties for various ranges of temperature and magnetic fields, and the optimal technology may be selected according to the application requirements. We briefly discuss the possible applications of MgB2 superconductors in devices, such as fault current limiters and electric machines.

2.
Microb Pathog ; 185: 106452, 2023 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-37972743

RESUMEN

The present investigation focuses on examining the clinical, histopathological, and ultrastructural changes that occurred in pig, during an outbreak of African swine fever (ASF) in 2022 in Assam, India. The disease initially manifested as a per-acute case with high mortality but without any evident clinical signs. Subsequently, some animals exhibited an acute form of the disease characterized by high fever (104-106 °F), anorexia, vomiting, respiratory distress, and bleeding from the anal and nasal orifices. During acute African swine fever virus (ASFV) infections, elevated levels of pro-inflammatory IL-1α, IL-1ß, IL-6, TNF, CCL2, CCL5, and CXCL10 were detected in the palatine tonsil, lymph nodes, spleen, and kidney using qPCR assay. These molecular changes were associated with haemorrhages, edemas, and lymphoid depletion. Postmortem examinations revealed prominent features such as splenomegaly with haemorrhages, haemorrhagic lymphadenitis, severe petechial haemorrhage in the kidney, pneumonia in the lungs, and necrotic palatine tonsil. Histopathological analysis demonstrated lymphocyte depletion in lymphoid organs, multi-organ haemorrhages, and interstitial pneumonia in the lungs. Scanning electron microscopy (SEM) further confirmed lymphocyte depletion in lymphoid organs through lymphocyte apoptosis and kidney damage with distorted tubules due to red blood cell destruction. Transmission electron microscopy reaffirmed lymphocyte apoptosis by observing chromatin condensation and nucleus margination in lymphocytes of lymphoid organs. These findings provide comprehensive insights into the clinical, histopathological, and ultrastructural aspects of ASF outbreak in pigs. Understanding the pathological changes associated with ASF can contribute to improved diagnosis, prevention, and control measures for this highly contagious and economically devastating viral disease.


Asunto(s)
Virus de la Fiebre Porcina Africana , Fiebre Porcina Africana , Porcinos , Animales , Fiebre Porcina Africana/epidemiología , Fiebre Porcina Africana/patología , Linfocitos , Brotes de Enfermedades , Hemorragia , Sus scrofa
3.
Biosci Biotechnol Biochem ; 87(10): 1102-1110, 2023 Sep 21.
Artículo en Inglés | MEDLINE | ID: mdl-37480241

RESUMEN

Proteins and peptides derived from various food sources are used in a variety of applications, including functional foods, pharmaceuticals, and cosmetics. The three-dimensional structure of proteins provides useful insights into their functions and essential information for the creation of proteins with new functions. In this review, a series of functional conversion technologies based on protein structural information derived from foods traditionally consumed in Japan, such as natto (fermented soybeans) and rice, are introduced. For natto, we first identified 2 types of Bacillus subtilis-derived endolytic and exolytic enzymes with different modes of action on soybean cell wall polysaccharides and then focused on the technology used to create an endolytic enzyme from an exolytic enzyme. By applying this technology, a method for creating novel bioactive peptides from rice seed proteins was established. The modified peptides created could provide diverse options for the production of substances such as pharmaceuticals and cosmetic materials.


Asunto(s)
Bacillus subtilis , Oryza , Pared Celular , Alimentos Funcionales , Japón , Glycine max , Preparaciones Farmacéuticas
4.
Int J Mol Sci ; 24(2)2023 Jan 11.
Artículo en Inglés | MEDLINE | ID: mdl-36674944

RESUMEN

DciA is the ancestral bacterial replicative helicase loader, punctually replaced during evolution by the DnaC/I loaders of phage origin. DnaC helps the helicase to load onto DNA by cracking open the hexameric ring, but the mechanism of loading by DciA remains unknown. We demonstrate by electron microscopy, nuclear magnetic resonance (NMR) spectroscopy, and biochemistry experiments that DciA, which folds into a KH-like domain, interacts with not only single-stranded but also double-stranded DNA, in an atypical mode. Some point mutations of the long α-helix 1 demonstrate its importance in the interaction of DciA for various DNA substrates mimicking single-stranded, double-stranded, and forked DNA. Some of these mutations also affect the loading of the helicase by DciA. We come to the hypothesis that DciA could be a DNA chaperone by intercalating itself between the two DNA strands to stabilize it. This work allows us to propose that the direct interaction of DciA with DNA could play a role in the loading mechanism of the helicase.


Asunto(s)
Proteínas de Escherichia coli , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/química , ADN Helicasas/metabolismo , ADN , Replicación del ADN , Bacterias/metabolismo , ADN de Cadena Simple , Proteínas Bacterianas/genética , Proteínas Bacterianas/química
5.
Angew Chem Int Ed Engl ; 60(20): 11201-11205, 2021 May 10.
Artículo en Inglés | MEDLINE | ID: mdl-33617079

RESUMEN

Chemical reduction of a naphthylene macrocycle, [6]cyclo-2,7-naphthylene ([6]CNAP, 1), with alkali metals, Li and K, revealed the accessibility of the doubly-reduced state of 1. The macrocyclic 12- anion was isolated in different coordination environments and crystallographically characterized. The single-crystal X-ray diffraction confirmed the formation of contact-ion complexes with one Li+ and two K+ ions in THF, and a "naked" dianion in the solvent-separated ion product with K+ ions in the presence of 18-crown-6 ether. The detailed structural analysis of 12- showed that the π-conjugation over the biaryl linkages between naphthylene panels were enhanced upon two-fold reduction, which was rationally explained by theoretical calculations.

6.
Bioorg Chem ; 106: 104502, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-33317841

RESUMEN

The new halogenated 1H-triazolo[4,5-b]pyridines and 1H-imidazo[4,5-b]pyridines were synthesised as analogues of known CK2 inhibitors: 4,5,6,7-tetrabromo-1H-benzotriazole (TBBt) and 4,5,6,7-tetrabromo-1H-benzimidazole (TBBi). Their influence on the activity of recombinant human CK2α, CK2α' and PIM1 kinases was determined. The most active inhibitors were di- and trihalogenated 1H-triazolo[4,5-b]pyridines (4a, 5a and 10a) with IC50 values 2.56, 3.82 and 3.26 µM respectively for CK2α. Furthermore, effect on viability of cancer cell lines MCF-7 (human breast adenocarcinoma) and CCRF-CEM (T lymphoblast leukemia) of all final compounds was evaluated. Finally, three crystal structures of complexes of CK2α1-335 with inhibitors 4a, 5a and 10a were obtained. In addition, new protocol was used to obtain high-resolution crystal structures of CK2α'Cys336Ser in complex with four inhibitors (4a, 5a, 5b, 10a).


Asunto(s)
Quinasa de la Caseína II/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Quinasa de la Caseína II/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Células MCF-7 , Estructura Molecular , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Relación Estructura-Actividad
7.
Food Sci Nutr ; 8(9): 4957-4964, 2020 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-32994957

RESUMEN

Several strains of microorganism are capable of converting carbohydrates into extracellular polysaccharide. The preset research is a first effort made to optimize extracellular polysaccharide (CRMEP) by Rhodotorula minuta ATCC 10658 using one factor at time and response surface methods. One factor at time was applied in the initial screening of substrates prior to optimization study. Of all the substrates examined, starch as carbon source and defatted soy bean powder as protein source were discovered to be best for CRMEP production. Response surface analysis revealed that 15 g/L starch and 30g/L defatted soy bean powder were the optimal chemical conditions. The model predicted 13.22 g/L for CRMEP, which went along with the experimentally observed result. Purification of CRMEP by anion-exchange column of DEAE-cellulose yielded RMEP. Structural investigation indicated that the main chain of RMEP was composed of (1 â†’ 3) and (1 â†’ 4)-linked mannopyranosyl residues, with branches attached to O-6 of some (1 â†’ 3)-linked mannopyranosyl residues. The branches were composed of Glcp-(1 â†’ residues.

8.
Angew Chem Int Ed Engl ; 59(37): 15923-15927, 2020 Sep 07.
Artículo en Inglés | MEDLINE | ID: mdl-32492236

RESUMEN

Chemical reduction of a benzo-fused double [7]helicene (1) with two alkali metals, K and Rb, provided access to three different reduced states of 1. The doubly-reduced helicene 12- has been characterized by single-crystal X-ray diffraction as a solvent-separated ion triplet with two potassium counterions. The triply- and tetra-reduced helicenes, 13- and 14- , have been crystallized together in an equimolar ratio and both form the contact-ion complexes with two Rb+ ions each, leaving three remaining Rb+ ions wrapped by crown ether and THF molecules. As structural consequence of the stepwise reduction of 1, the central axis of helicene becomes more compressed upon electron addition (1.42 Šin 14- vs. 2.09 Šin 1). This is accompanied by an extra core twist, as the peripheral dihedral angle increases from 16.5° in 1 to 20.7° in 14- . Theoretical calculations provided the pattern of negative charge build-up and distribution over the contorted helicene framework upon each electron addition, and the results are consistent with the X-ray crystallographic and NMR spectroscopic data.

9.
Int J Biol Macromol ; 124: 1321-1327, 2019 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-30244130

RESUMEN

IP3 is a ubiquitous second messenger in eukaryotic cells that triggers Ca2+-release from intracellular stores. IP3 binds to intracellular IP3-receptor (IP3R) and induces conformational change within the ligand-binding domain which regulates Ca2+-release; hence, both IP3 and IP3R are key components of the signal transduction mechanism. Here we present cDNA cloning of IP3-binding core (IBC) domain encoding only residues 224-604 of human IP3R type 2 that binds to IP3 with high affinity. RNA extraction, RT-PCR, PCR and cloning were carried out, and then the cloned DNA was checked by sequencing. Thereafter, expression vector pET-28a harboring the correct gene was transformed into different E. coli (DE3) strains and investigated its protein expression under various conditions. Finally, the IBC expression was induced at 20 °C for 20 h into BL21 strain at LB medium with 4 mM lactose and 0.5 mM IPTG, and then confirmed by western blotting. After protein purification, structural study was recorded in absence and presence of its ligand. Far-CD and intrinsic fluorescence spectra analysis of the purified protein with and without IP3 ligand showed change in secondary and tertiary IBC structure. Moreover, bioinformatics study demonstrated that the ligand binding site residues R269, K508 and R511 are conserved.


Asunto(s)
Receptores de Inositol 1,4,5-Trifosfato/química , Receptores de Inositol 1,4,5-Trifosfato/aislamiento & purificación , Inositol 1,4,5-Trifosfato/química , Secuencia de Aminoácidos , Animales , Sitios de Unión , Clonación Molecular , Biología Computacional , ADN Complementario/genética , ADN Complementario/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Expresión Génica , Vectores Genéticos/química , Vectores Genéticos/metabolismo , Humanos , Inositol 1,4,5-Trifosfato/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/genética , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Ligandos , Ratones , Modelos Moleculares , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Dominios y Motivos de Interacción de Proteínas , Estructura Terciaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Alineación de Secuencia , Homología de Secuencia de Aminoácido
10.
Carbohydr Polym ; 206: 220-228, 2019 Feb 15.
Artículo en Inglés | MEDLINE | ID: mdl-30553316

RESUMEN

A systematic structural characterization of dissolving grade cellulose pulp in aqueous NMMO solution is performed under the process conditions of lyocell slurry. Different types of cellulosic pulps such as hard/soft wood and acid sulfite/kraft sulphate pulps are used for the present study. The structural changes of pulp in lyocell slurry at different temperatures are characterized in terms of dimension, interstitial spaces, crystallinity using Optical (weight and thickness gain), SEM and XRD measurement technique, respectively. It was observed that kraft sulphate pulp has higher weight gain and lower thickness gain compared to acid sulphite pulps due to its pulping process chemistry. These results are further supported by SEM and XRD analysis. It is also found that above 50 °C, hardwood kraft sulphate pulp shows homogeneous and consistent swelling compared to other pulp combinations. These findings are commercially useful, because, homogeneous swelling of pulp is one of the essential parameters of slurry preparation.

11.
Open Access Maced J Med Sci ; 7(23): 4101-4109, 2019 Dec 15.
Artículo en Inglés | MEDLINE | ID: mdl-32165960

RESUMEN

BACKGROUND: Sofosbuvir (SOF) was approved in 2013 as a part of first-line treatment for hepatitis C virus (HCV); it has activity against all genotypes with extrahepatic adverse effects have recently arisen. AIM: Investigating sofosbuvir-induced alterations in the rat submandibular salivary gland (SMSG). METHODS: A group of 80 adult albino rats weighing about ± 150 gm were used in the experiment. The rats were divided into 3 groups: Group I (control group) received distilled water, Group II (experimental group) divided into 2 subgroups and received SOF 40 mg/kg/day dissolved in distilled water for 1 and 3 months and Group III (recovery group) allowed for 1 month of recovery after SOF withdrawal. All animals were sacrificed; the SMSG was dissected, and specimens were examined histologically and ultra-structurally. RESULTS: Compared to Group I, Group II subgroup (1) showed acinar and ductal vacuolisation, discontinuity of the epithelial lining associated with retained secretion and congested blood vessels. These changes were found to be exaggerated in the subgroup (2) accompanied by acinar and ductal shrinkage, interstitial oedema, haemorrhage, chronic inflammatory cells infiltration and loss of gland compactness. Amelioration of the histological changes was detected in Group III after SOF withdrawal. The ultrastructural examination confirmed these histological results. CONCLUSION: SOF had induced apparent alterations in the structure and ultrastructure of SMSG. The SOF-induced alterations were time-dependent, attributed mainly to mitochondrial toxicity and partially ameliorated by its withdrawal.

12.
J Mass Spectrom ; 53(12): 1230-1237, 2018 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-30286511

RESUMEN

We report herein a detailed structural study by collision-induced dissociation (CID) of nonglycosylated anthocyanins (anthocyanidins) using electrospray ionization triple quadrupole mass spectrometry (ESI-QqQ) and isotope labeling experiments to understand the fragmentation process often used in mass spectrometry analysis of this class of compounds. Tandem mass spectrometric product ion spectra for three anthocyanidins (cyanidin, delphynidin, and pelargonin) were evaluated to propose fragmentation mechanisms to this natural colorant class of organic compounds. The proposed rearrangements, retro Diels-Alder reaction, water loss, CO losses, and stable acylium ion formation, were evaluated based on tandem mass spectrometric experiments of normal and labeled precursor ions together to computational thermochemistry. B3LYP/6-311 + G** ab initio calculations studies were carried out to obtain energy diagrams to show the viability of the proposed mechanisms. The CO losses fragmentation channels have lower energies when compared with water losses and the other proposed fragmentations. The isotope labeling experiments indicate the H/D exchange of the hydroxyl protons and corroborate the proposed general fragmentation mechanism for anthocyanidins.

13.
Carbohydr Res ; 469: 23-30, 2018 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-30199787

RESUMEN

In recent years, 1,4-triazole rings are being widely used for the synthesis of carbohydrate derived biomimetics, due to their easy synthesis and wide range of functional group compatibility. These triazole rings lead to synthetic molecules with improved enzymatic stability, bioavailability, and structural diversity. In this present work, a benzoyl group has been introduced at the C-5 position of the triazole ring present in the synthetic glycoconjugates providing further structural diversity to the molecule. 5-Benzoyl 1,4-triazole ring containing glycoconjugates were synthesized using Cu(I) catalyzed [3 + 2] cycloaddition reaction of per-O-acetylated glycopyranosyl azide and phenyl acetylene followed by in situ electrophilic addition of benzoyl group to the Cu(I) coordinated triazole intermediate. The X-ray crystal structure of one of the 5-benzoyl 1,4-triazole linked glycoconjugate derived from d-xylose {1-N-(2,3,4-tri-O-acetyl-ß-d-xylopyranosyl)-4-phenyl-5-benzoyl-1,2,3-triazole} showed unique pattern of intermolecular CH…O interactions arranging the molecules in an anti-parallel orientation. The structure and morphology of the compounds were further explored using computational calculation and scanning electron microscopic (SEM) study which firmly established the uniqueness of 5-benzoyl 1,4-triazole linked glycoconjugates compared to that of 5-H 1,4-triazole linked derivative.


Asunto(s)
Glicoconjugados/química , Triazoles/química , Conformación de Carbohidratos , Catálisis , Modelos Moleculares
14.
Nanoscale Res Lett ; 13(1): 229, 2018 Aug 03.
Artículo en Inglés | MEDLINE | ID: mdl-30076473

RESUMEN

In this research, a facile co-precipitation method was used to synthesize pure and Mg-doped ZnO nanoparticles (NPs). The structure, morphology, chemical composition, and optical and antibacterial activity of the synthesized nanoparticles (NPs) were studied with respect to pure and Mg-doped ZnO concentrations (0-7.5 molar (M) %). X-ray diffraction pattern confirmed the presence of crystalline, hexagonal wurtzite phase of ZnO. Scanning electron microscope (SEM) images revealed that pure and Mg-doped ZnO NPs were in the nanoscale regime with hexagonal crystalline morphology around 30-110 nm. Optical characterization of the sample revealed that the band gap energy (Eg) decreased from 3.36 to 3.04 eV with an increase in Mg2+ doping concentration. Optical absorption spectrum of ZnO redshifted as the Mg concentration varied from 2.5 to 7.5 M. Photoluminescence (PL) spectra showed UV emission peak around 400 nm. Enhanced visible emission between 430 and 600 nm with Mg2+ doping indicated the defect density in ZnO by occupying Zn2+ vacancies with Mg2+ ions. Photocatalytic studies revealed that 7.5% Mg-doped ZnO NPs exhibited maximum degradation (78%) for Rhodamine B (RhB) dye under UV-Vis irradiation. Antibacterial studies were conducted using Gram-positive and Gram-negative bacteria. The results demonstrated that doping with Mg ions inside the ZnO matrix had enhanced the antibacterial activity against all types of bacteria and its performance was improved with successive increment in Mg ion concentration inside ZnO NPs.

15.
Bioorg Chem ; 80: 266-275, 2018 10.
Artículo en Inglés | MEDLINE | ID: mdl-29966873

RESUMEN

The new aminoalkyl-substituted derivatives of known CK2 inhibitors 4,5,6,7-tetrabromo-1H-benzimidazole (TBBi) and 4,5,6,7-tetrabromo-1H-benzotriazole (TBBt) were synthesized, and their influence on the activity of recombinant human CK2 α, CK2 holoenzyme and PIM1 kinases was evaluated. All derivatives inhibited the activity of studied kinases and the most efficient were aminopropyl-derivatives 8b and 14b. These compounds also exerted inhibition of cancer cell lines - CCRF-CEM (acute lymphoblastoid leukemia), MCF-7 (human breast cancer), and PC-3 (prostate cancer) proliferation and their EC50 is comparable with the value for clinically studied CK2 inhibitor CX-4945. Preliminary structure activity relationship analysis indicated that the spacer length affected antitumor potency, and two to three methylene units were more favorable. The complex of CK2 α1-335/8b was crystallized, both under high-salt conditions and under low-salt conditions giving crystals which diffracted X-rays to about 2.4 Šresolution, what enabled the determination of the corresponding 3D-structures.


Asunto(s)
Bencimidazoles/química , Quinasa de la Caseína II/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Triazoles/química , Bencimidazoles/metabolismo , Bencimidazoles/farmacología , Sitios de Unión , Quinasa de la Caseína II/genética , Quinasa de la Caseína II/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Humanos , Simulación de Dinámica Molecular , Fosforilación/efectos de los fármacos , Inhibidores de Proteínas Quinasas/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Estructura Terciaria de Proteína , Proteínas Proto-Oncogénicas c-pim-1/genética , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/química , Proteínas Recombinantes/aislamiento & purificación , Relación Estructura-Actividad , Triazoles/metabolismo , Triazoles/farmacología
16.
Int J Biol Macromol ; 107(Pt A): 1227-1234, 2018 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-28958818

RESUMEN

A new water-soluble polysaccharide named as AAPS-1, was extracted with warm water from the roots of Acanthophyllum acerosum and further purified by DEAE-cellulose A52 anion-exchange and Sephadex G-50 gel filtration chromatography. AAPS-1 was a neutral polysaccharide, with a molecular weight of 23.2kDa and a specific optical rotation of +181.3° (c 1.0, H2O), and comprised of Glucose, Galactose and Arabinose with a relative molar ratio of 1.6:5.1:1.0. Structure features of AAPS-1 were investigated by a combination of chemical and instrumental analysis, such as partial acid hydrolysis, methylation, periodate oxidation and Smith degradation, GC-MS, FTIR and NMR (13C and 1H). The data obtained indicate that AAPS-1 possessed a backbone of →6)-α-d-Galp-(1→ residues, with branches attached to O-2 (∼35%) by α-d-Glcp-(1→ and at O-3 (∼16%) by α-d-Galp-(1→ and by ß-l-Arap-(1→3)-ß-l-Arap-(1→3)-ß-l-Arap-(1→. The scavenging activity of AAPS-1 against DPPH radical was less than that of ascorbic acid at the same concentrations and the EC50 value of AAPS-1 was 1.4mg/ml.


Asunto(s)
Antioxidantes/química , Raíces de Plantas/química , Polisacáridos/química , Agua/química , Compuestos de Bifenilo/química , Caryophyllaceae/química , Carbohidratos de la Dieta , Depuradores de Radicales Libres/química , Galactosa/química , Glucosa/química , Hidrólisis , Espectroscopía de Resonancia Magnética , Estructura Molecular , Monosacáridos/química , Picratos/química , Espectroscopía Infrarroja por Transformada de Fourier
17.
Sci Adv ; 3(4): e1601601, 2017 04.
Artículo en Inglés | MEDLINE | ID: mdl-28435872

RESUMEN

The spontaneous formation of biological higher-order structures from smaller building blocks, called self-assembly, is a fundamental attribute of life. Although the protein self-assembly is a time-dependent process that occurs at the molecular level, its current understanding originates either from static structures of trapped intermediates or from modeling. Nuclear magnetic resonance (NMR) spectroscopy has the unique ability to monitor structural changes in real time; however, its size limitation and time-resolution constraints remain a challenge when studying the self-assembly of large biological particles. We report the application of methyl-specific isotopic labeling combined with relaxation-optimized NMR spectroscopy to overcome both size- and time-scale limitations. We report for the first time the self-assembly process of a half-megadalton protein complex that was monitored at the structural level, including the characterization of intermediate states, using a mutagenesis-free strategy. NMR was used to obtain individual kinetics data on the different transient intermediates and the formation of final native particle. In addition, complementary time-resolved electron microscopy and native mass spectrometry were used to characterize the low-resolution structures of oligomerization intermediates.


Asunto(s)
Proteínas Arqueales/química , Péptido Hidrolasas/química , Multimerización de Proteína , Pyrococcus horikoshii/enzimología , Resonancia Magnética Nuclear Biomolecular , Estructura Cuaternaria de Proteína
18.
Sci Adv ; 1(7): e1500734, 2015 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-26601243

RESUMEN

The structure of DNA was determined in 1953 by x-ray fiber diffraction. Several attempts have been made to obtain a direct image of DNA with alternative techniques. The direct image is intended to allow a quantitative evaluation of all relevant characteristic lengths present in a molecule. A direct image of DNA, which is different from diffraction in the reciprocal space, is difficult to obtain for two main reasons: the intrinsic very low contrast of the elements that form the molecule and the difficulty of preparing the sample while preserving its pristine shape and size. We show that through a preparation procedure compatible with the DNA physiological conditions, a direct image of a single suspended DNA molecule can be obtained. In the image, all relevant lengths of A-form DNA are measurable. A high-resolution transmission electron microscope that operates at 80 keV with an ultimate resolution of 1.5 Å was used for this experiment. Direct imaging of a single molecule can be used as a method to address biological problems that require knowledge at the single-molecule level, given that the average information obtained by x-ray diffraction of crystals or fibers is not sufficient for detailed structure determination, or when crystals cannot be obtained from biological molecules or are not sufficient in understanding multiple protein configurations.

19.
Int J Biol Macromol ; 81: 560-7, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26318664

RESUMEN

An insight into the effects of new ionic liquid-type gemini imidazolium cationic surfactants on the structure and function of the lipases is of prime importance for their potential application. Changes in the activity, stability and structure of Rhizopus oryzae lipase in the presence of novel gemini surfactants, [C16-3-C16im]Br2 and [C16-12-C16im]Br2 were probed in the present study. Surfactant with shorter spacer length, [C16-3-C16im]Br2 was found to be better in improving the hydrolytic activity and thermal stability of the lipase. For both the surfactants, activation was concentration dependent. CD spectroscopy results showed a decrease in α-helix and an increase in ß-sheet content in the presence of these surfactants. A higher structural change observed in presence of [C16-12-C16im]Br2 correlated with lower enzyme activity. Isothermal titration calorimetric studies showed the binding to be spontaneous in nature based on sequential two site binding model. The forces involved in binding were found to differ for the two surfactants proving that the spacer length is an important factor which governs the interaction. These surfactants could be used as promising components both in enzyme modification and media engineering for attaining the desired goals in biocatalytic reactions.


Asunto(s)
Calcitriol/análogos & derivados , Lipasa/química , Rhizopus/enzimología , Tensoactivos/química , Calcitriol/química , Calcitriol/farmacología , Catálisis , Activación Enzimática/efectos de los fármacos , Estabilidad de Enzimas , Hidrólisis , Estructura Molecular , Desplegamiento Proteico , Análisis Espectral , Tensoactivos/farmacología , Termodinámica
20.
Spectrochim Acta A Mol Biomol Spectrosc ; 134: 608-13, 2015 Jan 05.
Artículo en Inglés | MEDLINE | ID: mdl-25049172

RESUMEN

Structural, spectroscopic and thermal properties of SiO2-Al2O3-Sb2O3-Na2O glass system doped with 0.2 mol% Tm2O3 have been presented. Synthesis of antimony-silicate glasses with relatively low phonon energy (600 cm(-1), which implicates a small non-radiative decay rate) was performed by conventional high-temperature melt-quenching methods. The effect of SiO2/Sb2O3 ratio in fabricated Tm(3+) doped glass on thermal, structural and luminescence properties was investigated. On the basis of structural investigations decomposition of absorption bands in the infrared FTIR region was performed, thus determining that antimony ions are the only glass-forming ions, setting up the lattice of fabricated glasses. Luminescence band at the wavelength of 1.8 µm corresponding to (3)F4→(3)H6 transition in thulium ions was obtained under 795 nm laser pumping. It was observed that combination of relatively low phonon energy and greater separation of optically active centers in the fabricated glasses influenced in decreasing the luminescence intensity at 1800 nm.


Asunto(s)
Antimonio/química , Vidrio/química , Silicatos/química , Tulio/química , Mediciones Luminiscentes , Espectroscopía Infrarroja por Transformada de Fourier
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