共查询到20条相似文献,搜索用时 15 毫秒
1.
Electrochemically directed synthesis of oligonucleotides for DNA microarray fabrication 总被引:2,自引:0,他引:2
We demonstrate a new method for making oligonucleotide microarrays by synthesis in situ. The method uses conventional DNA synthesis chemistry with an electrochemical deblocking step. Acid is delivered to specific regions on a glass slide, thus allowing nucleotide addition only at chosen sites. The acid is produced by electrochemical oxidation controlled by an array of independent microelectrodes. Deblocking is complete in a few seconds, when competing side-product reactions are minimal. We demonstrate the successful synthesis of 17mers and discrimination of single base pair mismatched hybrids. Features generated in this study are 40 μm wide, with sharply defined edges. The synthetic technique may be applicable to fabrication of other molecular arrays. 相似文献
2.
We report the synthesis and the characterisation of the first electroactive ferrocene-labelled oligonucleotide phosphorothioate (ODN-Fc-Ps) probe obtained by automated synthesis. The grafting of ODN-Fc-Ps probe on gold electrode resulted in the appearance of the ferrocene redox couple in cyclic voltammetry confirming the effectiveness of the ODN grafting. The electrochemical response of the modified electrode was analysed in aqueous media before and after hybridisation with ODN target. The hybridisation with ODN target induces a large conformational change in the surface-confined DNA structure monitored by cyclic voltammetry of the ferrocene marker which confirms the potential of ferrocene-labelled oligonucleotide phosphorothioate to develop electrochemical DNA chips. 相似文献
3.
A cellular biosensing system for screening protein synthesis inhibitors has been developed by linking an electrochemical phosphate modulator and matrix-immobilized yeast cells with an optical sensing device. To screen the protein synthesis inhibitors, yeast phosphatase gene regulating system has been employed by linking an electrochemical phosphate modulator. Since the yeast phosphatase gene coding gammaAPase is expressed, when the phosphate concentration in solution is lowered below the threshold, the gammaAPase production is triggered by lowering the phosphate concentration with the electrochemical phosphate modulator, and monitored continuously with the photometric device. The electrochemical phosphate modulator was assembled with matrix-immobilized yeast cells. The module could insert to ordinal cuvette to monitor the induced gammaAPase activity in an ordinal photometer. Using the system, induction profile of protein synthesis was easily observed and was affected remarkably by various protein synthesis inhibitors. This seems promising that the system can be applied for first screening process of de novo protein inhibitors. The cellular biosensing system seems promising in screening protein synthesis inhibitors. 相似文献
4.
Mohammad Mazloum-Ardakani Nooshin Rajabzadeh Ali Benvidi Mohammad Mehdi Heidari 《Analytical biochemistry》2013
We have developed a simple and renewable electrochemical biosensor based on carbon paste electrode (CPE) for the detection of DNA synthesis and hybridization. CPE was modified with gold nanoparticles (AuNPs), which are helpful for immobilization of thiolated bioreceptors. AuNPs were characterized by scanning electron microscopy (SEM). Self-assembled monolayers (SAMs) of thiolated single-stranded DNA (SH–ssDNA) of the amelogenin gene was formed on CPE. The immobilization of the probe and its hybridization with the target DNA was optimized using different experimental conditions. The modified electrode was characterized by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). The electrochemical response of ssDNA hybridization and DNA synthesis was measured using differential pulse voltammetry (DPV) with methylene blue (MB) as an electroactive indicator. The new biosensor can distinguish between complementary and non-complementary strands of amelogenin ssDNA. Genomic DNA was extracted from blood and was detected based on changes in the MB reduction signal. These results demonstrated that the new biosensor could be used for sex determination. The proposed biosensor in this study could be used for detection and discrimination of polymerase chain reaction (PCR) products of amelogenin DNA. 相似文献
5.
During ATP synthesis, ATP synthase has to bind MgADP in the presence of an excess of MgATP. Thus, for efficient ATP synthesis it would be desirable if incoming substrate could be bound to a catalytic site with a preference for MgADP over MgATP. We tested three hypotheses predicting the existence of such a site. However, our results showed that, at least in absence of an electrochemical proton gradient, none of the three catalytic sites has a higher affinity for MgADP than for MgATP. 相似文献
6.
Meier T Morgner N Matthies D Pogoryelov D Keis S Cook GM Dimroth P Brutschy B 《Molecular microbiology》2007,65(5):1181-1192
Despite the thermodynamic problem imposed on alkaliphilic bacteria of synthesizing adenosine triphosphate (ATP) against a large inverted pH gradient and consequently a low electrochemical proton potential, these bacteria still utilize a proton-coupled F(1)F(o)-ATP synthase to synthesize ATP. One potential solution to this apparent thermodynamic problem would be the operation of a larger oligomeric c ring, which would raise the ion to ATP ratio, thus facilitating the conversion of a low electrochemical potential into a significant phosphorylation potential. To address this hypothesis, we have purified the oligomeric c ring from the thermoalkaliphilic bacterium Bacillus sp. strain TA2.A1 and determined the number of c-subunits using a novel mass spectrometry method, termed 'laser-induced liquid bead ion desorption' (LILBID). This technique allows the mass determination of non-covalently assembled, detergent-solubilized membrane protein complexes, and hence enables an accurate determination of c ring stoichiometries. We show that the Bacillus sp. strain TA2.A1 ATP synthase harbours a tridecameric c ring. The operation of a c ring with 13 subunits renders the thermodynamic problem of ATP synthesis at alkaline pH less severe and may represent a strategy for ATP synthesis at low electrochemical potential. 相似文献
7.
Starved whole cells of alkalophilic Bacillus firmus OF4 that are equilibrated at either pH 10.2, 9.5, or 8.5 synthesize ATP in response to a pH gradient that is imposed by rapid dilution of the cyanide-treated cells into buffer at pH 7.5. If a valinomycin-mediated potassium diffusion potential (positive out) is generated simultaneously with the pH gradient, then the rate of ATP synthesis and the level of synthesis achieved is much higher than upon imposition of a pH gradient alone. By contrast, imposition of a large chemical gradient of Na+, either in the presence or absence of a concomitant diffusion potential, fails to result in ATP synthesis. We conclude that this organism does not possess a sodium-motive ATPase that can be made to synthesize detectable levels of ATP by imposition of a suitably large chemical or electrochemical gradient of Na+. On the other hand, a proton-translocating ATPase is in evidence when protons are provided at very high pH, corroborating our earlier work on extremely alkalophilic bacilli. Oxidative phosphorylation must, then, be catalyzed in these organisms by a proton-translocating ATPase even though the putative bulk driving forces for such a catalyst are low under optimal growth conditions. Stable, imposed pH gradients of 1 unit, comparable to the magnitude of the total electrochemical proton gradient of growing cells, result in much lower ATP concentrations than observed in such cells. We hypothesize that ATP synthesis in growing cells utilizes protons that are made available by some localized pathway between proton pumps and the ATP synthase. 相似文献
8.
Diez M Zimmermann B Börsch M König M Schweinberger E Steigmiller S Reuter R Felekyan S Kudryavtsev V Seidel CA Gräber P 《Nature structural & molecular biology》2004,11(2):135-141
Synthesis of ATP from ADP and phosphate, catalyzed by F(0)F(1)-ATP synthases, is the most abundant physiological reaction in almost any cell. F(0)F(1)-ATP synthases are membrane-bound enzymes that use the energy derived from an electrochemical proton gradient for ATP formation. We incorporated double-labeled F(0)F(1)-ATP synthases from Escherichia coli into liposomes and measured single-molecule fluorescence resonance energy transfer (FRET) during ATP synthesis and hydrolysis. The gamma subunit rotates stepwise during proton transport-powered ATP synthesis, showing three distinct distances to the b subunits in repeating sequences. The average durations of these steps correspond to catalytic turnover times upon ATP synthesis as well as ATP hydrolysis. The direction of rotation during ATP synthesis is opposite to that of ATP hydrolysis. 相似文献
9.
The H(+)-ATPase from chloroplasts was brought into the active, reduced state. Then, an electrochemical potential difference of protons across the thylakoid membranes was generated by an acid-base transition, delta pH, combined with a K+/valinomycin diffusion potential, delta psi. The initial rate of ATP synthesis was measured with a rapid-mixing quenched-flow apparatus in the time-range between 20-150 ms. The rate of ATP synthesis depends in a sigmoidal way on delta pH. Increasing diffusion potentials shifts the delta pH-dependencies to lower delta pH values. Analysis of the data indicate that the rate of ATP synthesis depends on the electrochemical potential difference of protons irrespective of the relative contribution of delta pH and delta psi. 相似文献
10.
An electrochemical potential difference for protons (delta mu H+) across the membrane of bacterial chromatophores was induced by an artificially generated pH difference (delta pH) and a K+/valinomycin diffusion potential, delta phi. The initial rate of ATP synthesis was measured with a rapid-mixing quenched-flow apparatus in the time range between 70 ms and 30 s after the acid-base transition. The rate of ATP synthesis depends exponentially on delta pH. Increasing diffusion potentials shift the delta pH dependency to lower delta pH values. Diffusion potentials were calculated from the Goldman equation. Using estimated permeability coefficients, the rate of ATP synthesis depends only on the electrochemical potential difference of protons irrespective of the relative contribution of delta pH and delta phi. 相似文献
11.
H+/ATP stoichiometry of proton pump of turtle urinary bladder 总被引:2,自引:0,他引:2
Urinary acidification in the turtle urinary bladder is due to a reversible proton-translocating ATPase. To estimate the H+/ATP stoichiometry of this pump, we measured the delta G'ATP in the epithelial cells and the maximum e.m.f. generated by the pump. The latter is the maximal transepithelial electrochemical gradient for protons placed across the epithelium that is needed to nullify the rate of transport and averaged 179 +/- 7 mV. The delta G'ATP averaged 50.1 kJ/mol. The H+/ATP stoichiometry of these bladders was 2.92 +/- 0.1. In other experiments, the bladders were poisoned by iodoacetate and cyanide and a variable transepithelial electrochemical gradient for protons was placed across them. It was noted that ATP synthesis occurred at a transepithelial electrochemical gradient for protons greater than 120 mV. The delta G'ATP in other bladders treated identically averaged 40.0 kJ/mol, giving a H+/ATP stoichiometry of 3.4 +/- 0.1. We conclude that the H+/ATP stoichiometry of the proton pump of turtle urinary bladder is approximately 3. 相似文献
12.
We recently described the chemical synthesis of 3, 4-dihydroxyphenylacetaldehyde and 3,4-dihydroxyphenylglycolaldehyde, the monamine oxidase metabolites of dopamine and noradrenaline, respectively. We demonstrated the neurotoxicity of these compounds. Catecholamine nerve cells which synthesize these aldehydes die in degenerative brain diseases, such as Parkinson's and Alzheimer's. Here we describe a sensitive method for separating these catecholaldehydes from catecholamines and their other oxidative and methylated metabolites by microcolumn high-performance liquid chromatography with electrochemical detection. We then quantitate catecholamines and their major metabolites in human brain, plasma, and urine. The method can be used to determine the role of these catecholaldehydes in human disease. 相似文献
13.
Hydrolysis of urea by Ureaplasma urealyticum generates a transmembrane potential with resultant ATP synthesis. 总被引:6,自引:3,他引:3
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When urea is added to Ureaplasma urealyticum, it is hydrolysed internally by a cytosolic urease. Under our measuring conditions, and at an external pH of 6.0, urea hydrolysis caused an ammonia chemical potential equivalent to almost 80 mV and, simultaneously, an increase in proton electrochemical potential (delta p) of about 24 mV with resultant de novo ATP synthesis. Inhibition of the urease with the potent inhibitor flurofamide abolished both the chemical potential and the increase of delta p such that ATP synthesis was reduced to approximately 5% of normally obtained levels. Uncouplers of electrochemical gradients had little or no effect on these systems. The electrochemical parameters and ATP synthesis were measured similarly at three other external pH values. Any change in delta p was primarily via membrane potential (delta psi), and the level of de novo ATP synthesis was related to the increase in delta p generated upon addition of urea and more closely to the ammonia chemical potential. Although the organisms lack an effective mechanism for internal pH homeostasis, they maintained a constant delta pH. The data reported are consistent with, and give evidence for, the direct involvement of a chemiosmotic mechanism in the generation of around 95% of the ATP by this organism. Furthermore, the data suggest that the ATP-generating system is coupled to urea hydrolysis by the cytosolic urease via an ammonia chemical potential. 相似文献
14.
Developing advanced electrochemical energy storage and conversion (ESC) technologies based on renewable clean energy can alleviate severe global environmental pollution and energy crisis. The efficient preparation of functional electrode materials via a simple, green, and safe synthesis process is the key to the commercial feasibility of these ESC systems. Deep eutectic solvents (DESs) with easy-tunable solvent properties and recyclable features have emerged as novel solvent systems for designing and synthesizing various functional powder materials for ESC devices. In this paper, the application of DESs in the synthesis of energy-related functional powder materials is systematically reviewed. After briefly introducing the classification and synthesis of DESs, their critical roles in synthesizing powder materials are discussed. Then, the recent advances of DES-derived powder materials in ESC, including batteries, fuel cells, supercapacitors, and water splitting, are described in detail from the perspective of preparation-structure-activity. Finally, some challenges and development directions of the DESs-mediated synthesis of powder materials with high electrochemical performance for ESC applications are outlined. 相似文献
15.
Mixed metal sulfides (MMSs) have attracted increased attention as promising electrode materials for electrochemical energy storage and conversion systems including lithium‐ion batteries (LIBs), sodium‐ion batteries (SIBs), hybrid supercapacitors (HSCs), metal–air batteries (MABs), and water splitting. Compared with monometal sulfides, MMSs exhibit greatly enhanced electrochemical performance, which is largely originated from their higher electronic conductivity and richer redox reactions. In this review, recent progresses in the rational design and synthesis of diverse MMS‐based micro/nanostructures with controlled morphologies, sizes, and compositions for LIBs, SIBs, HSCs, MABs, and water splitting are summarized. In particular, nanostructuring, synthesis of nanocomposites with carbonaceous materials and fabrication of 3D MMS‐based electrodes are demonstrated to be three effective approaches for improving the electrochemical performance of MMS‐based electrode materials. Furthermore, some potential challenges as well as prospects are discussed to further advance the development of MMS‐based electrode materials for next‐generation electrochemical energy storage and conversion systems. 相似文献
16.
The monoamine oxidase A metabolite of noradrenaline, 3,4-dihydroxyphenylglycolaldehyde, is the precursor of 3,4-dihydroxymandelic acid, and 3,4-dihydroxyphenylglycol, metabolites of noradrenaline. Owing to difficulties in purifying this aldehyde, it has not been previously characterized or identified in biological sources. This paper describes an enzymatic synthesis, purification, and characterization of 3,4-dihydroxyphenylglycolaldehyde. The aldehyde metabolite is identified in postmortem human brain using high-performance liquid chromatography and electrochemical detection. We estimate the concentration in human hippocampus to be 0.164 +/- 0.05 nmol/g. The importance of this aldehyde metabolite of noradrenaline is discussed. 相似文献
17.
Ultrasensitive DNA sensor based on gold nanoparticles/reduced graphene oxide/glassy carbon electrode
Ali Benvidi Afsaneh Dehghani Firouzabadi Seyed Mohammad Moshtaghiun Mohammad Mazloum-Ardakani Marzieh Dehghan Tezerjani 《Analytical biochemistry》2015
We have designed a simple and novel electrochemical biosensor based on glassy carbon electrode (GCE) for DNA detection. GCE was modified with reduced graphene oxide (RGO) and gold nanoparticles (AuNPs) by the electrochemical method, which is helpful for immobilization of thiolated bioreceptors. The electrode modification processes were characterized by scanning electron microscopy (SEM) and electrochemical methods. Then a single-stranded DNA (ssDNA) probe for BRCA1 5382 insC mutation detection was immobilized on the modified electrode for a specific time. The experimental conditions, such as probe immobilization time and target DNA (complementary DNA) hybridization time and temperature with probe DNA, were optimized using electrochemical methods. The electrochemical response for DNA hybridization and synthesis was measured using electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) methods. The calibration graph contains two linear ranges; the first part is in the range of 3.0 × 10−20 to 1.0 × 10−12 M, and the second segment part is in the range of 1.0 × 10−12 to 1.0 × 10−7 M. The biosensor showed excellent selectivity for the detection of the complementary sequences from noncomplementary sequences, so it can be used for detection of breast cancer. 相似文献
18.
《Neurochemistry international》1986,8(2):165-170
The synthesis of noradrenaline was measured, using high-performance liquid chromatography with electrochemical detection, in synaptosomal fractions prepared from rat hippocampus. Noradrenaline synthesis is depressed by adenosine deaminase and the adenosine antagonist, 8-phenyltheophylline and stimulated by the adenosine agonist, 2-chloroadenosine. β-Adrenoceptor stimulation also increases synthesis. The adenosine receptors and β-adrenoceptors do not interact. Both receptor-mediated effects are distinct from, and additive with, the acceleration of synthesis by potassium-depolarisation.The results are compatible with an adenosine-receptor and β-adrenoceptor stimulation of adenylate cyclase, leading to a cyclic AMP-mediated activation of tyrosine hydroxylase. 相似文献
19.
Sakthivel Kandaiah 《Inorganica chimica acta》2009,362(11):4009-4012
We present the electrochemical synthesis by galvanostatic electro-oxidation method and single crystal structure determination of a complex of silver(II) with a macrocyclic N donor ligand crystallized with hexafluorophosphate anions. The crystal structure analysis showed a penta-coordinated environment of silver(II) and displacement of the metal centre from the rectangular planar configuration of the macrocyclic N donor which is due to axial coordination by a solvent acetonitrile molecule. Both PF6− anions were modelled with two-fold disorder. 相似文献
20.
P Sharma SK Tuteja V Bhalla G Shekhawat VP Dravid CR Suri 《Biosensors & bioelectronics》2013,39(1):99-105
We report a novel in-situ electrochemical synthesis approach for the formation of functionalized graphene-graphene oxide (fG-GO) nanocomposite on screen-printed electrodes (SPE). Electrochemically controlled nanocomposite film formation was studied by transmission electron microscopy (TEM) and Raman spectroscopy. Further insight into the nanocomposite has been accomplished by the Fourier transformed infrared spectroscopy (FTIR), thermal gravimetric analysis (TGA) and X-ray diffraction (XRD) spectroscopy. Configured as a highly responsive screen-printed immunosensor, the fG-GO nanocomposite on SPE exhibits electrical and chemical synergies of the nano-hybrid functional construct by combining good electronic properties of functionalized graphene (fG) and the facile chemical functionality of graphene oxide (GO) for compatible bio-interface development using specific anti-diuron antibody. The enhanced electrical properties of nanocomposite biofilm demonstrated a significant increase in electrochemical signal response in a competitive inhibition immunoassay format for diuron detection, promising its potential applicability for ultra-sensitive detection of range of target analytes. 相似文献