Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 14 de 14
Filtrar
1.
J Am Chem Soc ; 145(22): 11939-11944, 2023 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-37224473

RESUMO

Organometallic reagents are effective for carbon-carbon bond formation; however, consumption of stoichiometric amounts of metals is problematic. We developed electrochemical allylation reactions of imines catalyzed by nitrogen-doped carbon-supported single-atom zinc, which were fixed on a cathode to afford a range of homoallylic amines efficiently. The system could suppress generation of metallic waste, and the catalyst electrode showed advantages over bulk zinc in terms of activity and robustness. An electrochemical flow reaction was also successfully performed to produce the homoallylic amine continuously with minimum amounts of waste.

2.
Langmuir ; 32(15): 3665-9, 2016 Apr 19.
Artigo em Inglês | MEDLINE | ID: mdl-27027321

RESUMO

Macroscopic continuous hierarchical ordering of achiral nanotube "imogolite" was achieved by thixotropic gelation of imogolite with chiral hydroxy acid and their flow-orienting/subsequent standing for uniaxial alignments of imogolite. The chirality change of the hydroxy acids resulted in an inversion of the helical ordering. The study presented here first exhibits the millimeter-scale supramolecular chirality induced by angstrom-scale molecular handedness in the architecture of nanotubes.

3.
J Biol Chem ; 289(52): 35686-94, 2014 Dec 26.
Artigo em Inglês | MEDLINE | ID: mdl-25294880

RESUMO

VanX is a d-alanyl-d-alanine (d-Ala-d-Ala) dipeptidase encoded in the vancomycin-resistance vanA gene cluster. Here we report that strong bacteriolysis occurred when isolated VanX was expressed in Escherichia coli at temperatures lower than 30 °C, which was unexpected because the vanA operon confers vancomycin resistance by protecting the cell wall. Therefore, we monitored cell lysis by measuring sample turbidity with absorbance at 590 nm and VanX expression using SDS-PAGE. No cell lysis was observed when VanX was expressed, even in large quantities, in the cell inclusion bodies at 37 °C, suggesting that a natively folded VanX is required for lysis. In addition, VanX mutants with suppressed dipeptidase activity did not lyse E. coli cells, confirming that bacteriolysis originated from the dipeptidase activity of VanX. We also observed shape changes in E. coli cells undergoing VanX-mediated lysis with optical microscopy and classified these changes into three classes: bursting, deformation, and leaking fluid. Optical microscopic image analysis fully corroborated our interpretation of the turbidity changes in the samples. From a practical perspective, the finding that VanX expressed in isolation induces cell lysis suggests that inhibitors of VanA and VanH that act downstream from VanX could provide a new class of therapeutic chemicals against bacteria expressing the vancomycin-resistance gene cluster.


Assuntos
Proteínas de Bactérias/genética , Bacteriólise/genética , D-Ala-D-Ala Carboxipeptidase Tipo Serina/genética , Proteínas de Bactérias/biossíntese , Proteínas de Bactérias/química , Escherichia coli/enzimologia , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/ultraestrutura , Genes Bacterianos , Família Multigênica , Estrutura Secundária de Proteína , D-Ala-D-Ala Carboxipeptidase Tipo Serina/biossíntese , D-Ala-D-Ala Carboxipeptidase Tipo Serina/química , Resistência a Vancomicina/genética
4.
Chemistry ; 21(47): 17091-9, 2015 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-26427615

RESUMO

Novel ordered lamellar mesostructure pZrPS-2 was hydrothermally prepared by using zirconium propoxide and 4-(EtO)2OPC6H4Si(OEt)3 (pPPS-E), which was hydrolyzed to organic building units substituted with both siliconate and phosphonate groups, in the presence of Cn TAB and TMAOH. The pZrPS-2 materials were obtained at a Zr/PPS ratio of 2 or higher and the basal spacing was increased by using a longer-chain surfactant (n = 12-18). Removal of the occluded surfactants at 300 °C resulted in retention of the lamellar structure with negligible shrinkage of the interlayer distance. Nitrogen adsorption studies revealed the ordered mesoporous nature of pZrPS-2 with a pore diameter of approximately 2 to 3 nm. The lamellar structure is assumed to be composed of layers that include zirconia-based crystalline nanodomains and interlayer pillars mainly based on PPS units. Although lamellar structures with the same crystalline phase also formed when no surfactant was added or when the meta isomer of PPS was used, no mesoporous materials were obtained except pZrPS-2. A possible schematic model to elucidate these results is also proposed.

5.
Small ; 10(9): 1813-20, 2014 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-24573908

RESUMO

In this paper, we describe stimuli-responsive hydrogels prepared from a rigid rod-like polyelectrolyte 'imogolite' and a dicarboxylic acid. The hydrogel exhibited thixotropy in response to mechanical shock within the order of seconds or sub-seconds. Here, using the latest structural/rheological characterisation techniques, the relationship between the structural transition processes and the shear thinning was estimated. The evidence obtained by the experiments revealed for the first time the direct relationship between the microscopic structural change and the macroscopic thixotropic behavior that have been extensively discussed. The thixotropic hydrogel has the hierarchical architecture in the combination of imogolite and dicarboxylic acid, i.e., sheathed nanotubes/hydroclusters of cross-bridged nanotubes/frameworks. The formation and disintegration of the network structure upon resting and agitating, respectively, were the origin of gel/sol transition (thixotropy), although the hydroclusters of cross-bridged nanotubes were maintained throughout the transition.

6.
Anal Biochem ; 439(2): 212-7, 2013 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-23624113

RESUMO

Escherichia coli is a versatile, low-cost, and popular host for expressing recombinant proteins. However, extracting recombinant proteins from E. coli requires cell wall breakage, which is both time- and effort-consuming. Here we report a novel cell breakage method based on our recent finding that VanX, which is a d-Ala-d-Ala dipeptidase encoded in a vancomycin-resistant VanA gene cluster, exhibits a strong cell lysis activity when expressed in isolation in E. coli. In our strategy, we coexpress VanX with the target protein, causing cell autolysis and release of the cellular content into the culture medium. We demonstrated this strategy for two model proteins, a green fluorescent protein variant (GFPuv) and Gaussia luciferase, and optimized the autolysis conditions and coexpression vectors. The fluorescence activity of GFPuv collected from the medium was identical to that of GFPuv purified by conventional methods. Cell breakage by VanX-mediated autolysis is very simple to implement and will efficiently complement traditional methods.


Assuntos
Proteínas de Bactérias/metabolismo , Bacteriólise/efeitos dos fármacos , Escherichia coli/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Luciferases/metabolismo , D-Ala-D-Ala Carboxipeptidase Tipo Serina/metabolismo , Proteínas de Bactérias/genética , Meios de Cultura , Escherichia coli/química , Regulação Bacteriana da Expressão Gênica/fisiologia , Regulação Enzimológica da Expressão Gênica , Proteínas de Fluorescência Verde/química , Luciferases/química , D-Ala-D-Ala Carboxipeptidase Tipo Serina/genética
7.
Micromachines (Basel) ; 12(9)2021 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-34577778

RESUMO

Solid-state nanopores are widely used as a platform for stochastic nanopore sensing because they can provide better robustness, controllable pore size, and higher integrability than biological nanopores. However, the fabrication procedures, including thin film preparation and nanopore formation, require advanced micro-and nano-fabrication techniques. Here, we describe the simple fabrication of solid-state nanopores in a commercially available material: a flat thin carbon film-coated micro-grid for a transmission electron microscope (TEM). We attempted two general methods for nanopore fabrication in the carbon film. The first method was a scanning TEM (STEM) electron beam method. Nanopores were fabricated by irradiating a focused electron beam on the carbon membrane on micro-grids, resulting in the production of nanopores with pore diameters ranging from 2 to 135 nm. The second attempt was a dielectric breakdown method. In this method, nanopores were fabricated by applying a transmembrane voltage of 10 or 30 V through the carbon film on micro-grids. As a result, nanopores with pore diameters ranging from 3.7 to 1345 nm were obtained. Since these nanopores were successfully fabricated in the commercially available carbon thin film using readily available devices, we believe that these solid-state nanopores offer great utility in the field of nanopore research.

8.
Chem Asian J ; 16(3): 232-236, 2021 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-33336531

RESUMO

Although strong acid-treated metal oxides are useful heterogeneous superacid catalysts for various organic transformations, they usually have a limited density of acidic sites due to their low surface areas. Herein, heterogeneous trifluoromethanesulfonic acid immobilized nitrogen-doped carbon-incarcerated titanium nanoparticle (NP) catalysts have been developed that are composed of well-dispersed, small Ti NPs (ca 7 nm) that are otherwise difficult to achieve using acid-treated metal oxides. The catalysts showed high activity for Friedel-Crafts acylation with low titanium loading (2 mol%, <1 mg of metal for 1 mmol of substrate). A range of microscopic, spectroscopic and physicochemical studies revealed that the nitrogen-doped carbon immobilized the trifluoromethanesulfonic acid and that the addition of metals further changed the nature of the acidic species and enhanced catalytic activity.

9.
Artigo em Inglês | WPRIM | ID: wpr-826308

RESUMO

BACKGROUND@#The Japanese health and welfare industry has a shortage of professional caregivers, and work-related accidents and injuries among this group are therefore especially critical issues. This study aimed to examine the factors associated with turnover intentions and work-related injuries and accidents among professional caregivers in Japan.@*METHODS@#Self-report questionnaires were distributed to care workers (N = 1396) at 26 geriatric-care facilities. The questionnaire addressed basic attributes, work and organizational characteristics, wage adequacy, and intrinsic motivations for work (e.g., "being suited to caring work"). Social-relational aspects of the work environment were assessed via three subscales of the Social Capital and Ethical Climate in the Workplace instrument (i.e., "Social Capital in the Workplace," "Exclusive Workplace Climate," and "Ethical Leadership"). Dependent variables were the experience of work-related accidents or injuries in the prior year and organizational and occupational turnover intentions. We used datasets of professional caregivers for analyses.@*RESULTS@#The response rate was 68% (N = 949). Among the 667 professional caregivers, 63% were female. On multivariable logistic regression analysis for work-related accidents and injuries for each sex, those with higher scores for "being suited to caring work" were found to experience significantly fewer work-related accidents and injuries (odds ratio [OR] = 0.78, p < 0.01) among female caregivers. Male caregivers who perceived an exclusive workplace climate experienced more work-related accidents and injuries (OR = 1.61, p < 0.01). However, experience of work-related accidents and injuries did not show significant relationships with organizational and occupational turnover intentions. Additionally, "being suited to caring work" (OR = 0.73, p < 0.01) and ethical leadership (OR = 0.76, p < 0.05) were found to be negatively associated with organizational turnover intentions. "Being suited to caring work" (OR = 0.61, p < 0.01), inadequacy of wage (OR = 2.22, p < 0.05), and marital status (OR = 2.69, p < 0.01) were also associated with occupational turnover intentions of professional caregivers.@*CONCLUSIONS@#These findings highlight the need to foster intrinsic motivations for work as well as providing a supportive and ethical work environment to reduce high turnover rates and work-related injuries and accidents among professional caregivers.


Assuntos
Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Acidentes de Trabalho , Atitude do Pessoal de Saúde , Cuidadores , Psicologia , Intenção , Japão , Satisfação no Emprego , Traumatismos Ocupacionais , Reorganização de Recursos Humanos , Autorrelato
10.
Biomaterials ; 31(1): 58-66, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19836832

RESUMO

We developed a hydrogel self-assembling method driven by the interaction between recombinant tax-interactive protein-1 (TIP1) with the PDZ domain in a molecule, which is fused to each end of the triangular trimeric CutA protein (CutA-TIP1), and a PDZ domain-recognizable peptide which is covalently bound to each terminus of four-armed poly(ethylene glycol) (PDZ-peptide-PEG). Genetic manipulation based on molecular-dynamic simulation generated a cell-adhesive RGD tripeptidyl sequence in the CutA loop region [CutA(RGD)-TIP1]. Spontaneous viscoelastic hydrogel formation occurred when either CutA-TIP1- or CutA(RGD)-TIP1-containing buffer solution and PDZ-peptide-PEG-containing buffer solutions were stoichiometrically mixed. Dynamic viscoelasticity measurement revealed shear stress-dependent reversible-phase transformation: a spontaneous viscoelastic hydrogel was formed at low shear stress, but it was transformed into a sol at high shear stress. Upon the cessation of shear, hydrogel was restored. When chondrocytes were pre-mixed with one of these two components containing buffer solutions, the stoichiometric mixed solution was also spontaneously gelled. Individual rounded cells and multicellular aggregates were entrapped within both hydrogels without substantial cellular impairment regardless of the presence or absence of RGD motif in the CutA-TIP1 molecule. The potential use of such a shear-sensitive hydrogel for injectable cell delivery into diseased or lost cartilage tissue is discussed.


Assuntos
Condrócitos/citologia , Hidrogéis , Oligopeptídeos/química , Proteínas/química , Sequência de Bases , Células Cultivadas , Primers do DNA , Eletroforese em Gel de Poliacrilamida , Humanos , Microscopia Eletrônica de Varredura , Modelos Moleculares , Simulação de Dinâmica Molecular
11.
Protein Sci ; 18(5): 960-9, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19384998

RESUMO

Self-assembly of artificially designed proteins is extremely desirable for nanomaterials. Here we show a novel strategy for the creation of self-assembling proteins, named "Nanolego." Nanolego consists of "structural elements" of a structurally stable symmetrical homo-oligomeric protein and "binding elements," which are multiple heterointeraction proteins with relatively weak affinity. We have established two key technologies for Nanolego, a stabilization method and a method for terminating the self-assembly process. The stabilization method is mediated by disulfide bonds between Cysteine-residues incorporated into the binding elements, and the termination method uses "capping Nanolegos," in which some of the binding elements in the Nanolego are absent for the self-assembled ends. With these technologies, we successfully constructed timing-controlled and size-regulated filament-shape complexes via Nanolego self-assembly. The Nanolego concept and these technologies should pave the way for regulated nanoarchitecture using designed proteins.


Assuntos
Nanoestruturas , Nanotecnologia/métodos , Engenharia de Proteínas , Multimerização Proteica , Proteínas , Animais , Simulação por Computador , Dissulfetos/metabolismo , Humanos , Camundongos , Modelos Moleculares , Nanoestruturas/química , Nanoestruturas/ultraestrutura , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/ultraestrutura , Ligação Proteica , Estabilidade Proteica , Proteínas/química , Proteínas/metabolismo , Proteínas/ultraestrutura , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/ultraestrutura
12.
Langmuir ; 23(5): 2668-73, 2007 Feb 27.
Artigo em Inglês | MEDLINE | ID: mdl-17269804

RESUMO

To characterize the molecular basis of specific interactions of PDZ proteins, dynamic force spectroscopy (DFS) for the PDZ protein Tax-interacting protein-1 (TIP-1) and its recognition peptide (PDZ-pep) derived from beta-catenin was performed using an atomic force microscope (AFM), together with measurement of thermodynamic and kinetic parameters using surface plasmon resonance (SPR). The unbinding force of this pair was measured under different conditions of AFM tip-retraction velocity. The relationship between the unbinding force and the logarithmic force-loading rate, that is, the dynamic force spectrum, exhibited two different rate regimes, for each of which the forces increased linearly with the force-loading rate. On the basis of the theoretical treatment of the Bell-Evans model, the positions of two different activation barriers in the reaction coordinate and dissociation rate constants in each barrier were evaluated from slopes and x-intercepts of the two linear regimes (first barrier: 0.04 nm and 1.10 x 10 s(-1); second barrier: 0.21 nm and 2.77 x 10(-2) s(-1), respectively). Although two-step unbinding kinetics between TIP-1 and PDZ-pep was suggested from the DFS analysis, SPR results showed single-step dissociation kinetics with a rate constant of 2.89 x 10(-1) s(-1). Different shapes of the free energy profile of the unbinding process were deduced from each result of DFS and SPR. The reason for such topographic differences in the energy landscape is discussed in relation to the differences in the pathways of forced unbinding and spontaneous dissociation.


Assuntos
Proteínas de Transporte/química , Físico-Química/métodos , Peptídeos/química , Análise Espectral/métodos , Vidro , Glutationa Transferase/química , Cinética , Ligantes , Metiltransferases , Microscopia de Força Atômica/métodos , Ligação Proteica , Estrutura Terciária de Proteína , Ressonância de Plasmônio de Superfície , Propriedades de Superfície , Termodinâmica
13.
Biomacromolecules ; 6(5): 2776-84, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-16153118

RESUMO

To determine the intermolecular force on protein-protein interaction (PPI) by atomic force microscopy (AFM), a photograft-polymer spacer for protein molecules on both surfaces of the substrate and AFM probe tip was developed, and its effectiveness was assessed in a PPI model of a pair of human serum albumin (HSA) and its monoclonal antibody (anti-HSA). A carboxylated photoiniferter, N-(dithiocarboxy)sarcosine, was derivatized on both surfaces of the glass substrate and AFM probe tip, and subsequently water-soluble nonionic vinyl monomers, N,N-dimethylacrylamide (DMAAm), were graft-polymerized on them upon ultraviolet light irradiation. DMAAm-photograft-polymerized spacers with carboxyl groups at the growing chain end but with different chain lengths on both surfaces were prepared. The proteins were covalently bound to the carboxyl terminus of the photograft-polymer chain using a water-soluble condensation agent. The effects of the graft-spacer length on the profile of the force-distance curves and on the unbinding characteristics (unbinding force and unbinding distance) were examined in comparison with those in the case of the commercially available poly(ethylene glycol) (PEG) spacer. The frequency of the nonspecific adhesion force profile was markedly decreased with the use of the photograft spacers. Among the force curves detected, a high frequency of single-peak curves indicating the unbinding process of a single pair of proteins and a very low frequency of multiple-peak profiles were observed for the photograft spacers, regardless of the graft chain length, whereas a high frequency of no-force peaks was noted. These observations were in marked contrast with those for the PEG spacer. The force peak values determined ranged from 88 to 94 pN, irrespective of the type of spacer, while the standard deviation of force distribution observed for the photograft spacer was lower than that for the PEG spacer, indicating that the photograft spacers provide a higher accuracy of force determination.


Assuntos
Anticorpos/química , Reações Antígeno-Anticorpo , Antígenos/química , Microscopia de Força Atômica/métodos , Adesividade , Anticorpos Monoclonais/química , Materiais Biocompatíveis/química , Vidro , Humanos , Luz , Polietilenoglicóis/química , Polímeros/química , Proteínas/química , Sarcosina/análogos & derivados , Sarcosina/química , Albumina Sérica/química , Estresse Mecânico , Propriedades de Superfície , Tiocarbamatos/química , Aderências Teciduais
14.
Cytotechnology ; 40(1-3): 139-49, 2002 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19003114

RESUMO

Reactive oxygen species (ROS) cause irreversible damage to biological macromolecules, resulting in many diseases. Reduced water (RW) such as hydrogen-rich electrolyzed reduced water and natural reduced waters like Hita Tenryosui water in Japan and Nordenau water in Germany that are known to improve various diseases, could protect a hamster pancreatic beta cell line, HIT-T15 from alloxan-induced cell damage. Alloxan, a diabetogenic compound, is used to induce type 1 diabetes mellitus in animals. Its diabetogenic effect is exerted via the production of ROS. Alloxan-treated HIT-T15 cells exhibited lowered viability, increased intracellular ROS levels, elevated cytosolic free Ca(2+) concentration, DNA fragmentation, decreased intracellular ATP levels and lowering of glucose-stimulated release of insulin. RW completely prevented the generation of alloxan-induced ROS, increase of cytosolic Ca(2+) concentration, decrease of intracellular ATP level, and lowering of glucose-stimulated insulin release, and strongly blocked DNA fragmentation, partially suppressing the lowering of viability of alloxan-treated cells. Intracellular ATP levels and glucose-stimulated insulin secretion were increased by RW to 2-3.5 times and 2-4 times, respectively, suggesting that RW enhances the glucose-sensitivity and glucose response of beta-cells. The protective activity of RW was stable at 4 degrees C for over a month, but was lost by autoclaving. These results suggest that RW protects pancreatic beta-cells from alloxan-induced cell damage by preventing alloxan-derived ROS generation. RW may be useful in preventing alloxan-induced type 1-diabetes mellitus.

SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA