Direct electron transfer Dotaz Zobrazit nápovědu
At present direct electron transfer between an electrode and protein ranks among promising areas of biosensor research. This review focuses on current trends and methods in the development of third-generation amperometric biosensors. It describes the construction of the biosensors of three generations and enzyme immobilization techniques. The design of the electrode interface and biosensor architecture are essential in bioelectrocatalysis, fast electron transfer and electrode construction. Differences in the architectures usually reflect specific electron transfer mechanisms. In the second-generation biosensors, the preferred design consists of biosensors with mediators tightly bound in the active layer of the electrode surface. Attempts at the direct electron transfer employ techniques like self-assembled monolayers, conducting polymers, reconstitution of the prosthetic group and apoenzyme in active holoenzyme and integration of nanotechnology. Despite considerable efforts, no commercially available biosensors based on the direct electron transfer have been developed so far. Modern approaches such as redox proteins and enzymes with engineered electron pathways can make this type of biosensors commercially attractive.
Three-dimensional (3D) printing technology offers attractive possibilities for many fields. In electrochemistry, 3D printing technology has been used to fabricate customized 3D-printed electrodes as a platform to develop bio/sensing, energy generation and storage devices. Here, we use a 3D-printed graphene/polylactic (PLA) electrode made by additive manufacturing technology and immobilize horseradish peroxidase (HRP) to create a direct electron transfer enzyme-based biosensors for hydrogen peroxide detection. Gold nanoparticles are included in the system to confirm and facilitate heterogeneous electron transfer. This work opens a new direction for the fabrication of third-generation electrochemical biosensors using 3D printing technology, with implications for applications in the environmental and biomedical fields.
Alphaproteobacteria, which are the most abundant microorganisms of temperate oceans, produce phage-like particles called gene transfer agents (GTAs) that mediate lateral gene exchange. However, the mechanism by which GTAs deliver DNA into cells is unknown. Here we present the structure of the GTA of Rhodobacter capsulatus (RcGTA) and describe the conformational changes required for its DNA ejection. The structure of RcGTA resembles that of a tailed phage, but it has an oblate head shortened in the direction of the tail axis, which limits its packaging capacity to less than 4,500 base pairs of linear double-stranded DNA. The tail channel of RcGTA contains a trimer of proteins that possess features of both tape measure proteins of long-tailed phages from the family Siphoviridae and tail needle proteins of short-tailed phages from the family Podoviridae. The opening of a constriction within the RcGTA baseplate enables the ejection of DNA into bacterial periplasm.
- MeSH
- bakteriofágy genetika fyziologie ultrastruktura MeSH
- DNA bakterií genetika MeSH
- elektronová kryomikroskopie MeSH
- přenos genů horizontální MeSH
- regulace genové exprese u bakterií MeSH
- Rhodobacter capsulatus genetika virologie MeSH
- Siphoviridae genetika fyziologie ultrastruktura MeSH
- technika přenosu genů * MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
INTRODUCTION: Propofol causes a profound inhibition of fatty acid oxidation and reduces spare electron transfer chain capacity in a range of human and rodent cells and tissues-a feature that might be related to the pathogenesis of Propofol Infusion Syndrome. We aimed to explore the mechanism of propofol-induced alteration of bioenergetic pathways by describing its kinetic characteristics. METHODS: We obtained samples of skeletal and cardiac muscle from Wistar rat (n = 3) and human subjects: vastus lateralis from hip surgery patients (n = 11) and myocardium from brain-dead organ donors (n = 10). We assessed mitochondrial functional indices using standard SUIT protocol and high resolution respirometry in fresh tissue homogenates with or without short-term exposure to a range of propofol concentration (2.5-100 μg/ml). After finding concentrations of propofol causing partial inhibition of a particular pathways, we used that concentration to construct kinetic curves by plotting oxygen flux against substrate concentration during its stepwise titration in the presence or absence of propofol. By spectrophotometry we also measured the influence of the same propofol concentrations on the activity of isolated respiratory complexes. RESULTS: We found that human muscle and cardiac tissues are more sensitive to propofol-mediated inhibition of bioenergetic pathways than rat's tissue. In human homogenates, palmitoyl carnitine-driven respiration was inhibited at much lower concentrations of propofol than that required for a reduction of electron transfer chain capacity, suggesting FAO inhibition mechanism different from downstream limitation or carnitine-palmitoyl transferase-1 inhibition. Inhibition of Complex I was characterised by more marked reduction of Vmax, in keeping with non-competitive nature of the inhibition and the pattern was similar to the inhibition of Complex II or electron transfer chain capacity. There was neither inhibition of Complex IV nor increased leak through inner mitochondrial membrane with up to 100 μg/ml of propofol. If measured in isolation by spectrophotometry, propofol 10 μg/ml did not affect the activity of any respiratory complexes. CONCLUSION: In human skeletal and heart muscle homogenates, propofol in concentrations that are achieved in propofol-anaesthetized patients, causes a direct inhibition of fatty acid oxidation, in addition to inhibiting flux of electrons through inner mitochondrial membrane. The inhibition is more marked in human as compared to rodent tissues.
- MeSH
- druhová specificita MeSH
- krysa rodu rattus MeSH
- lidé středního věku MeSH
- lidé MeSH
- mastné kyseliny metabolismus MeSH
- oxidace-redukce účinky léků MeSH
- potkani Wistar MeSH
- propofol farmakologie MeSH
- respirační komplex I metabolismus MeSH
- respirační komplex IV metabolismus MeSH
- senioři MeSH
- srdeční mitochondrie metabolismus MeSH
- svalové mitochondrie metabolismus MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
BACKGROUND: Conversely to common coated iron oxide nanoparticles, novel naked surface active maghemite nanoparticles (SAMNs) can covalently bind DNA. Plasmid (pDNA) harboring the coding gene for GFP was directly chemisorbed onto SAMNs, leading to a novel DNA nanovector (SAMN@pDNA). The spontaneous internalization of SAMN@pDNA into cells was compared with an extensively studied fluorescent SAMN derivative (SAMN@RITC). Moreover, the transfection efficiency of SAMN@pDNA was evaluated and explained by computational model. METHODS: SAMN@pDNA was prepared and characterized by spectroscopic and computational methods, and molecular dynamic simulation. The size and hydrodynamic properties of SAMN@pDNA and SAMN@RITC were studied by electron transmission microscopy, light scattering and zeta-potential. The two nanomaterials were tested by confocal scanning microscopy on equine peripheral blood-derived mesenchymal stem cells (ePB-MSCs) and GFP expression by SAMN@pDNA was determined. RESULTS: Nanomaterials characterized by similar hydrodynamic properties were successfully internalized and stored into mesenchymal stem cells. Transfection by SAMN@pDNA occurred and GFP expression was higher than lipofectamine procedure, even in the absence of an external magnetic field. A computational model clarified that transfection efficiency can be ascribed to DNA availability inside cells. CONCLUSIONS: Direct covalent binding of DNA on naked magnetic nanoparticles led to an extremely robust gene delivery tool. Hydrodynamic and chemical-physical properties of SAMN@pDNA were responsible of the successful uptake by cells and of the efficiency of GFP gene transfection. GENERAL SIGNIFICANCE: SAMNs are characterized by colloidal stability, excellent cell uptake, persistence in the host cells, low toxicity and are proposed as novel intelligent DNA nanovectors for efficient cell transfection.
- MeSH
- biofyzikální jevy MeSH
- DNA chemie genetika MeSH
- genetické vektory MeSH
- koloidy chemie MeSH
- lidé MeSH
- nanočástice chemie ultrastruktura MeSH
- plazmidy chemie genetika MeSH
- technika přenosu genů * MeSH
- transfekce metody MeSH
- transmisní elektronová mikroskopie MeSH
- velikost částic MeSH
- železité sloučeniny chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
As is the case with batch-based tableting processes, continuous tablet manufacturing can be conducted by direct compression or with a granulation step such as dry or wet granulation included in the production procedure. In this work, continuous manufacturing tests were performed with a commercial tablet formulation, while maintaining its original material composition. Challenges were encountered with the feeding performance of the API during initial tests which required designing different powder pre-blend compositions. After the pre-blend optimization phase, granules were prepared with a roller compactor. Tableting was conducted with the granules and an additional brief continuous direct compression run was completed with some ungranulated mixture. The tablets were assessed with off-line tests, applying the quality requirements demanded for the batch-manufactured product. Chemical maps were obtained by Raman mapping and elemental maps by scanning electron microscopy with energy-dispersive X-ray spectroscopy. Large variations in both tablet weights and breaking forces were observed in all tested samples, resulting in significant quality complications. It was suspected that the API tended to adhere to the process equipment, accounting for the low API content in the powder mixture and tablets. These results suggest that this API or the tablet composition was unsuitable for manufacturing in a continuous line; further testing could be continued with different materials and changes in the process.
During translation termination in bacteria, the release factors RF1 and RF2 are recycled from the ribosome by RF3. While high-resolution structures of the individual termination factors on the ribosome exist, direct structural insight into how RF3 mediates dissociation of the decoding RFs has been lacking. Here we have used the Apidaecin 137 peptide to trap RF1 together with RF3 on the ribosome and visualize an ensemble of termination intermediates using cryo-electron microscopy. Binding of RF3 to the ribosome induces small subunit (SSU) rotation and swivelling of the head, yielding intermediate states with shifted P-site tRNAs and RF1 conformations. RF3 does not directly eject RF1 from the ribosome, but rather induces full rotation of the SSU that indirectly dislodges RF1 from its binding site. SSU rotation is coupled to the accommodation of the GTPase domain of RF3 on the large subunit (LSU), thereby promoting GTP hydrolysis and dissociation of RF3 from the ribosome.
- MeSH
- elektronová kryomikroskopie MeSH
- Escherichia coli genetika metabolismus MeSH
- GTP-fosfohydrolasy metabolismus MeSH
- kationické antimikrobiální peptidy farmakologie MeSH
- konformace proteinů MeSH
- peptidy - faktory ukončení metabolismus MeSH
- proteiny z Escherichia coli metabolismus MeSH
- proteosyntéza účinky léků MeSH
- ribozomální proteiny metabolismus MeSH
- ribozomy metabolismus MeSH
- RNA transferová metabolismus MeSH
- simulace molekulového dockingu MeSH
- terminace translace peptidového řetězce MeSH
- vazba proteinů MeSH
- vazebná místa MeSH
- velké podjednotky ribozomu bakteriální metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Článek je pokračováním pojednání (I) o technice, resp. bioinženýrských metodách pro praxia výzkum v dermatovenerologii. V tomto II. oddíle jsou probrány vyšetřovací metody určené k zobrazováníkůže a jejích struktur. Lze je rozdělit na přímé a nepřímé. V první části přímých metod jevěnována pozornost klasické fotografii a nové digitální fotografii. Jsou porovnány vlastnosti, technickémožnosti a přednosti obou způsobů, kvalita a využití. Probrány jsou způsoby archivace,počítačového zpracování, komprimace dat.Zmíněny jsou i způsoby makro- a mikrofotografií, včetně stereomikroskopie. Mezi nové metodypatří i videomikroskopie a její provádění v polarizovaném světle.Tato elektronická mikroskopie poskytuje při spojení s příslušným softwarem možnost ještěkvalitnějšího zpracování dat, především kožních nádorů (melanomů) a névů. Počítačové posuzovánína principu tzv. neuronových sítí poskytuje ještě kvalitnější ABCD+E+T analýzu s vyšší pravděpodobnostízáchytu melanomu. Systém je využitelný i pro ostatní uchování a popis obrazů. Přípojtlačítkem„video-mail“ dává možnost použití v teledermatologii, tj. při teledermatoskopii, teledermatopatologii,při konzultacích a edukaci.V další části jsou uvedeny mikroskopické metody, jako jsou konfokální a elektronová (skenovací)mikroskopie s jejich konstrukčními a technickými parametry a využitím.Do stati přímých metod patří ještě kapilaroskopie a fluorescenční videomikroskopie, optickákoherentní tomografie kůže a protonová magnetická rezonance kůže.K nepřímým metodám zařazujeme dopplerometrii (A-, B-, C-, M-mode scan), laser-Doppler-průtokometrii,měření nerovnosti kožního povrchu (profilometrii) a kožní viziometr. Jsou opět uvedenyjejich konstrukční základy, s technickým a praktickým použitím.
The article is a continuation of part I dealing with the technique and bioengineering methods forpractice and research in dermatovenerology. In part II the authors discuss examination methods forimaging of the skin and its structures. They can be divided into direct and indirect ones. In the firstpart of direct methods attention is paid to classical photography and new digital photography. Theauthors compare properties, technical possibilities and advantages of both methods, their qualityand application. They discuss also methods of archivation, computer processing and comprimationof data.The authors mention methods of macro- and microphotography, incl. stereomicroscopy. Newmethods include also videomicroscopy incl. its implementation in polarized light.This electronic microscopy provides when combined with the appropriate software the possibilityof high standard processing of data, in particular skin tumours (melanomas) and naevi. Computer-assisted evaluation on the principle of so-called neuron networks makes even higher-standardABCD+E+T analysis possible with a greater probability of melanoma detection. The system can beused also for other preservation and description of images. Connection by button „video-mail“ makesit possible to use of in teledermatology, i.e. in teledermatoscopy, teledermatopathology, duringconsultations and education. In the subsequent part microscopic methods are presented such as confocal and electron(scanning) microscopy with their design, technical parameters and use.The section on direct methods includes also capillaroscopy and fluorescent videomicroscopy,optic coherent tomography of the skin and proton magnetic resonance of the skin.Indirect methods include dopplerometry (A-, B-, C-, M-mode scan), laser-doppler flowmetry,assessment of inequalities of the skin surface (profilometry) and skin visiometry. Their design,technical and practical use are described.
- MeSH
- dermatologie metody MeSH
- elektronová mikroskopie metody využití MeSH
- konfokální mikroskopie metody využití MeSH
- lidé MeSH
- magnetická rezonanční tomografie metody využití MeSH
- ultrasonografie dopplerovská metody využití MeSH
- videomikroskopie metody využití MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- přehledy MeSH
Objective: The electronic patient record (EPR) has been introduced into nursing homes with the aim of reducing time spent on documentation, improving documentation quality and increasing transferability of information, all of which should facilitate care provision. However, previous research has shown that EPR may be creating new burdens for staff. The purpose of this literature review is to explore how EPR is facilitating or hindering care provision in nursing homes. Methods: An integrative literature review was carried out using four electronic databases to search for relevant articles. After screening, 22 articles were included for thematic synthesis. Results: Thematic synthesis resulted in six analytical themes linked to care provision: time for direct care; accountability; assessment and care planning; exchange of information; risk awareness; and person-centered care. Conclusion: For EPR to facilitate care provision in nursing homes, consideration should be given to the type of device used for documentation, as well as the types of applications, the functionality, content, and structure of EPR. Further research exploring the experiences of end users is required to identify the optimal characteristics of an EPR system specifically for use in nursing homes.
Electrochemical methods can be used not only for the sensitive analysis of proteins but also for deeper research into their structure, transport functions (transfer of electrons and protons), and sensing their interactions with soft and solid surfaces. Last but not least, electrochemical tools are useful for investigating the effect of an electric field on protein structure, the direct application of electrochemical methods for controlling protein function, or the micromanipulation of supramolecular protein structures. There are many experimental arrangements (modalities), from the classic configuration that works with an electrochemical cell to miniaturized electrochemical sensors and microchip platforms. The support of computational chemistry methods which appropriately complement the interpretation framework of experimental results is also important. This text describes recent directions in electrochemical methods for the determination of proteins and briefly summarizes available methodologies for the selective labeling of proteins using redox-active probes. Attention is also paid to the theoretical aspects of electron transport and the effect of an external electric field on the structure of selected proteins. Instead of providing a comprehensive overview, we aim to highlight areas of interest that have not been summarized recently, but, at the same time, represent current trends in the field.
