Immobilization of Ferrocene-Modified SNAP-Fusion Proteins

The supramolecular assembly of proteins on surfaces has been investigated via the site-selective incorporation of a supramolecular moiety on proteins. To this end, fluorescent proteins have been site-selectively labeled with ferrocenes, as supramolecular guest moieties, via SNAP-tag technology. The assembly of guest-functionalized SNAP-fusion proteins on cyclodextrin- and cucurbit [7]uril-coated surfaces yielded stable monolayers. The binding of all ferrocene fusion proteins is specific as determined by surface plasmon resonance. Micropatterns of the fusion proteins, on patterned cyclodextrin and cucurbituril surfaces, have been visualized using fluorescence microscopy. The SNAP-fusion proteins were also immobilized on cyclodextrin vesicles. The supramolecular SNAP-tag labeling of proteins, thus, allows for the assembly of modified proteins via supramolecular host-guest interaction on different surfaces in a controlled manner. These findings extend the toolbox of fabricating supramolecular protein patterns on surfaces taking advantage of the high labeling efficiency of the SNAP-tag with versatile supramolecular moieties.     

Synthesis and Characterization of Mesoporous Silica Functionalized with Calix[]arene Derivatives

This work reports a new method to covalently attach calix[4]arene derivatives onto MCM-41, using a diisocyanate as a linker. The modified mesoporous silicates were characterized by fourier transform infrared spectroscopy (FTIR), thermal analysis (TGA) and elemental analysis. The FTIR spectra and TGA analysis verified that the calix[4]arene derivates are covalently attached to the mesoporous silica. The preservation of the MCM-41 channel system was checked by X-ray diffraction and nitrogen adsorption analysis.     

In Vitro Treatment of Melanoma Brain Metastasis by Simultaneously Targeting the MAPK and PI3K Signaling Pathways

Malignant melanoma is the most lethal form of skin cancer, with a high propensity to metastasize to the brain. More than 60% of melanomas have the BRAF^V600E mutation, which activates the mitogen-activated protein kinase (MAPK) pathway [1]. In addition, increased PI3K (phosphoinositide 3-kinase) pathway activity has been demonstrated, through the loss of activity of the tumor suppressor gene, PTEN [2]. Here, we treated two melanoma brain metastasis cell lines, H1_DL2, harboring a BRAF^V600E mutation and PTEN loss, and H3, harboring WT (wild-type) BRAF and PTEN loss, with the MAPK (BRAF) inhibitor vemurafenib and the PI3K pathway associated mTOR inhibitor temsirolimus. Combined use of the drugs inhibited tumor cell growth and proliferation in vitro in H1_DL2 cells, compared to single drug treatment. Treatment was less effective in the H3 cells. Furthermore, a strong inhibitory effect on the viability of H1_DL2 cells, when grown as 3D multicellular spheroids, was seen. The treatment inhibited the expression of pERK1/2 and reduced the expression of pAKT and p-mTOR in H1_DL2 cells, confirming that the MAPK and PI3K pathways were inhibited after drug treatment. Microarray experiments followed by principal component analysis (PCA) mapping showed distinct gene clustering after treatment, and cell cycle checkpoint regulators were affected. Global gene analysis indicated that functions related to cell survival and invasion were influenced by combined treatment. In conclusion, we demonstrate for the first time that combined therapy with vemurafenib and temsirolimus is effective on melanoma brain metastasis cells in vitro. The presented results highlight the potential of combined treatment to overcome treatment resistance that may develop after vemurafenib treatment of melanomas.     

Contribution of Natural Inhibitors to the Understanding of the PI3K/PDK1/PKB Pathway in the Insulin-mediated Intracellular Signaling Cascade

The critical initial steps in insulin action include phosphorylation of adapter proteins and activation of phosphatidylinositol 3-kinase (PI3K). One of important components in this process is a protein called Akt/protein kinase B (PKB). The work of numerous different researchers indicates a role of PKB in regulating insulin-stimulated glucose uptake. The crucial role of lipid second messengers in PKB activation has been dissected through the use of the PI3K-specific inhibitors wortmannin and {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002. Receptor-activated PI3K synthesizes the lipid second messenger PtdIns[3,4,5]-trisphosphate, leading to the recruitment of PKB to the membrane. Membrane attachment of PKB is mediated by its pleckstrin homology domain binding to PtdIns[3,4,5]-trisphosphate or PtdIns[3,4]-bisphosphate with high affinity. Activation of PKB alpha is then achieved at the plasma membrane by phosphorylation of Thr308 in the activation-loop of the kinase domain and Ser473 in the carboxy-terminal regulatory region, respectively. 3-Phosphoinositide-dependent protein kinase-1 (PDK1) is responsible for T308 phosphorylation. The usage of specific inhibitors and natural compound has significantly contributed to investigate the molecular mechanism of PI3K/PDK1/PKB signaling pathway, leading to the putative therapeutics benefits of patients. This review focuses on the contribution of natural inhibitor or compound in our understanding of the mechanism by which insulin induces, especially in PI3K/PDK1/PKB signaling.     

不同链长的烷基硫醇自组装膜在不同腐蚀介质中的抗腐蚀性阻抗研究

文章通过在铜表面上制备高质量的不同链长的烷基硫醇自组装膜并研究烷基硫醇自组装膜在不同腐蚀介质NaCl、HCl、H2SO4中的抗腐蚀能力,从而对其抗腐蚀性能进行比较研究。结果表明:铜电极上组装的C18SH、C12SH、C6SHSAMs在不同的腐蚀介质中对基底均有相当好的腐蚀保护作用;在其它条件相同的情况下,烷基硫醇的碳链越长,自组装膜对铜基底的腐蚀保护能力越强。     

Characterization of a Low Shrinkage Dental Composite Containing Bismethylene Spiroorthocarbonate Expanding Monomer

In this study, a novel dental composite based on the unsaturated bismethylene spiroorthocarbonate expanding monomer 3,9-dimethylene-1,3,5,7-tetraoxa-spiro[5,5]undecane (BMSOC) and bisphenol-S-bis(3-meth acrylate-2-hydroxypropyl)ether (BisS-GMA) was prepared. CQ (camphorquinone) of 1 wt % and DMAEMA (2-(dimethylamino)ethyl methacrylate) of 2 wt % were used in a photoinitiation system to initiate the copolymerization of the matrix resins. Distilled water contact angle measurements were performed for the wettability measurement. Degree of conversion, volumetric shrinkage, contraction stress and compressive strength were measured using Fourier Transformation Infrared-FTIR spectroscopy, the AccuVol and a universal testing machine, respectively. Within the limitations of this study, it can be concluded that the resin composites modified by bismethylene spiroorthocarbonate and BisS-GMA showed a low volumetric shrinkage at 1.25% and a higher contact angle. The lower contraction stress, higher degree of conversion and compressive strength of the novel dental composites were also observed.     

Analogies Between Digital Radio and Chemical Orthogonality as a Method for Enhanced Analysis of Molecular Recognition Events

Acoustic wave biosensors are a real-time, label-free biosensor technology, which have been exploited for the detection of proteins and cells. One of the conventional biosensor approaches involves the immobilization of a monolayer of antibodies onto the surface of the acoustic wave device for the detection of a specific analyte. The method described within includes at least two immobilizations of two different antibodies onto the surfaces of two separate acoustic wave devices for the detection of several analogous analytes. The chemical specificity of the molecular recognition event is achieved by virtue of the extremely high (nM to pM) binding affinity between the antibody and its antigen. In a standard ELISA (Enzyme-Linked ImmunoSorbent Assay) test, there are multiple steps and the end result is a measure of what is bound so tightly that it does not wash away easily. The fact that this “gold standard” is very much not real time, masks the dance that is the molecular recognition event. X-Ray Crystallographer, Ian Wilson, demonstrated more than a decade ago that antibodies undergo conformational change during a binding event[1, 2]. Further, it is known in the arena of immunochemistry that some antibodies exhibit significant cross-reactivity and this is widely termed antibody promiscuity. A third piece of the puzzle that we will exploit in our system of acoustic wave biosensors is the notion of chemical orthogonality. These three biochemical constructs, the dance, antibody promiscuity and chemical orthogonality will be combined in this paper with the notions of in-phase (I) and quadrature (Q) signals from digital radio to manifest an approach to molecular recognition that allows a level of discrimination and analysis unobtainable without the aggregate. As an example we present experimental data on the detection of TNT, RDX, C4, ammonium nitrate and musk oil from a system of antibody-coated acoustic wave sensors.     

Electrochemical investigation of dynamic interfacial processes at 1-octadecanethiol-modified copper electrodes in halide-containing solutions

1-Octadecanethiol (C18SH) monolayers were self-assembled on the fresh and active copper surface pretreated by nitric acid etching method. The surface properties of the alkanethiol-modified copper electrode in halide-containing solutions were characterized systematically by using several electrochemical methods, including polarization curves, cyclic voltammetry, electrochemical impedance spectroscopy (EIS) and electrochemical noise (EN). The results show that C18SH self-assembled monolayers (SAMs) onto copper provide a flexible method that can protect the underlying copper against corrosion. With the immersion time of SAMs-coated copper electrode in NaCl and HCl corrosive solutions increasing, a slow loss of corrosion protection ability of SAMs indicates dynamic processes occurring at the electrode/solution interface and in the monolayers, such as expansion of the defects and transport of corrosive ions through defects of SAMs. Electrochemical noise (EN) is employed to detect the alkanethiol-modified copper surfaces immersed in HCl solution. This observation suggests the pitting process associate with dynamic processes in the 1-octadecanethiol layer.     

Synthesis of Analogues of Gingerol and Shogaol,the Active Pungent Principles from the Rhizomes of Zingiber officinale and Evaluation of Their Anti-Platelet Aggregation Effects

The present study was aimed at discovering novel biologically active compounds based on the skeletons of gingerol and shogaol, the pungent principles from the rhizomes of Zingiber officinale. Therefore, eight groups of analogues were synthesized and examined for their inhibitory activities of platelet aggregation induced by arachidonic acid, collagen, platelet activating factor, and thrombin. Among the tested compounds, [6]-paradol (5b) exhibited the most significant anti-platelet aggregation activity. It was the most potent candidate, which could be used in further investigation to explore new drug leads.     

Thermodynamic and kinetic aspects of the electron transfer reaction of bovine cytochrome c immobilized on 4-mercaptopyridine and 11-mercapto-1-undecanoic acid films

Bovine cytochrome c (cyt c) was adsorbed on a polycrystalline gold electrode coated with 4-mercaptopyridine and 11-mercapto-1-undecanoic acid self-assembled monolayers (SAMs) and the thermodynamics and kinetics of the heterogeneous protein-electrode electron transfer (ET) reaction were determined by cyclic voltammetry. The E°′ values for the immobilized protein were found to be lower than those for the corresponding diffusing species. The thermodynamic parameters for protein reduction ( and ) indicate that the stabilization of the ferric state due to protein–SAM interaction is enthalpic in origin. The kinetic data suggest that a tunneling mechanism is involved in the ET reaction: the distance between the redox center of the protein and the electrode surface can be efficiently evaluated using the Marcus equation.     

Electrochemical impedance and solid-state electrical characterization of silicon (1 1 1) modified with ω-functionalized alkyl monolayers

In order to reveal the relationship between the dipole moment and electric properties of organically modified semiconductor, a series of ω-functionalized alkyl monolayers on oxide-free silicon (Si-(CH₂)₁₀COOH, Si-(CH₂)₁₁CH₃, Si-(CH₂)₁₀COOC₂H₅, and Si-(CH₂)₁₁OH) was prepared and characterized. Electrochemical impedance measurements showed excellent insulating effects of these monolayers. It has been demonstrated that the dielectric constants of these organic monolayers are affected by the dipole moments introduced by the different terminal-groups. Solid-state electrical measurements showed that mercury | ω-functionalized alkyl monolayer | silicon junctions exhibit molecular tunability and a clear correlation between ideality factor and the film thickness/dielectric constant ratio. The barrier height is approximately proportional to the dipole moment of the monolayer. These results augment the possibility of fine-tuning the electrical properties of silicon-based microelectronic devices using functionalized organic monolayers.     

Stability of horseradish peroxidase immobilized on different cinnamic carbohydrate esters

Four types of organic polymer were used to cover 3 mm diameter glass beads for subsequent use in immobilizing horseradish peroxidase (HRP). These supports were the totally cinnamoylated derivatives of D ‐glucosone (GSOCN), D ‐sorbitol (SOTCN), ethyl‐D ‐glucopyranoside (EGSCN) or inuline (INCN). In some assays, a partially cinnamoylated derivative, 2,3,4,6‐tetracinnamoyl‐D ‐glucopyranoside (GPSTCN) (obtained from the EGSCN derivative by hydrolysis in acidic medium) was used. Polymerization and cross‐linking of the derivatives obtained initially was carried out by irradiation in the ultraviolet region, where these polymers show the greatest degree of sensitivity. The enzyme was immobilized by adsorption to the support. Immobilized enzymatic activity varied with the incubation time used (2–21 h), depending on the chemical nature of the immobilization support. The affinity of HRP for H₂O₂ and 2,2′‐azinobis(3‐ethylbenzothiazolinesulfonic acid) (ABTS) was slightly lower in the case of the enzyme immobilized on the GSOCN, SOTCN and EGSCN derivatives than in the case of the soluble enzyme, as can be seen from the corresponding apparent kinetic constants ( and ). However, the affinity of HRP immobilized on the INCN derivative for these substrates was higher than that shown by the enzyme in solution. In turn, the enzyme immobilized on all the supports was more resistant than the soluble form to inactivation by H₂O₂ and by heat at neutral pH. When the stability and durability of the immobilized HRP were analysed, the cinnamoylated derivatives prepared were seen to function as suitable supports for immobilizing peroxidase and to be suitable for industrial application of the enzyme. Copyright © 2004 Society of Chemical Industry     

Electrochemical impedance spectroscopy of carboxylic-acid terminal alkanethiol self assembled monolayers on GaAs substrates

Adsorbate-induced charge depolarization can influence the organization of self assembled monolayers (SAMs) on semiconductor surfaces, especially as a function of the SAM functional group, SAM length and substrate dopant level and type. Based on systematic differences in the frequency response of the electrochemical impedance and phase data for carboxylic acid (COOH) terminal alkanethiol monolayers of varying alkane chain length assembled on GaAs substrates of different dopant level and type, we assessed the relative monolayer quality through fits to an appropriate equivalent circuit analog to compare the proportion of defects and SAM-induced semiconductor depolarization. At the open circuit potential in the NaCl-phosphate buffer, while SAMs on p+ GaAs were of marginally better quality than those on p GaAs, SAMs on n+ GaAs exhibited a far superior quality than those formed on n GaAs. COOH-terminal SAMs of longer chain lengths formed higher quality monolayers at all the substrate doping levels. COOH-terminal SAM modified n and n+ GaAs surfaces were passivated and stable over a wider voltage range than SAM modified p and p+ GaAs surfaces, from cyclic voltammetry. The poorer quality of COOH-terminal SAMs formed on GaAs substrates at the lower doping levels is attributed to the disorder as a result of the enhanced degree of charge depolarization at these surfaces, as substantiated by systematic variations in the space charge capacitance upon SAM modification that suggest a negative surface dipole.     

Nano-Silica Catalyzed Synthesis,Solvent Effect,NMR Spectral and DFT Studies of Some Imidazole Derivatives

A series of novel imidazole derivatives has been designed and synthesized using nano-SiO₂ as an efficient catalyst. Synthesized compounds have been characterized by ¹H and ¹³C-NMR spectral studies. The significant features of this nanocatalyst are high product yield, short reaction times and a vast range of substrates usage. Proton and ¹³C chemical shifts of the synthesized compounds were calculated. The absorption and emission properties of imidazole derivatives were studied in several solvents. Polar solvents favor the stabilization of excitation of the imidazole derivative. Decrease in the total dipole moment of the solvent molecules (non-polar solvents) results in the change of the molecular charge distributions of the imidazole derivatives. Optimization of 4,5-dimethyl-2-phenyl-1-m-tolyl-1H-imidazole (1) was performed by DFT at B3LYP/6-31G (d, p) using Gaussian-03.     

Protection of self-assembled monolayers formed from triethyl phosphate and mixed self-assembled monolayers from triethyl phosphate and cetyltrimethyl ammonium bromide for copper against corrosion

Self-assembled monolayers (SAMs) of triethyl phosphate (TEP) and the mixed SAMs of TEP with cetyltrimethyl ammonium bromide (CTAB) were prepared on copper surfaces. The protection abilities of these films against copper corrosion in 0.2 M NaCl aqueous solution were investigated using electrochemical impedance spectroscopy (EIS) and polarization techniques. Results showed that the inhibition efficiency (IE) increased with an increase in the immersion time of copper in the TEP-containing solutions. When the TEP films were modified with CTAB, the ability of the corrosion inhibition of the mixed films improved markedly. Furthermore, the influence of an external magnetic field on the formation of SAMs was studied. Ab initio calculations were performed to provide some theoretical studies for our experiment.     

Self-Assembled Monolayers of Vinyltriethoxysilane and Vinyltrichlorosilane Deposited on Silicon Dioxide Surfaces

Abstract

This study was aimed at deposition of self-assembled monolayers (SAMs) using vinyltriethoxysilane (VTES) and vinyltrichlorosilane (VTCS) molecules chemisorbed on silicon dioxide surfaces. The kinetics of SAM formation on planar glass substrates and silicon wafers was characterized by contact angle measurements. The surface free energy and its dispersion and polar components enabled to estimate the time of immersion required to deposit compact SAMs. Adsorption of organosilane molecules as a function of immersion time was characterized by X-ray photoelectron spectroscopy. The SAM thickness was evaluated by spectroscopic ellipsometry. Surface topography of deposited layers was investigated by atomic force microscopy (AFM). The VTCS/glass combination exhibited the fastest kinetics but the deposit was not uniform and included local agglomerates. The hydrophobic vinyl groups at deposit surface resulted in a surface free energy of 32 mJ/m².     

硅烷自组装单分子层生长条件的优化

对长碳链硅烷在玻璃片表面自组装形成单分子层膜进行研究。以甲苯为溶剂,使辛烷基三乙氧基硅烷(OTES)、十二烷基三甲氧基硅烷(DTMS)、3-胺丙基三乙氧基硅烷(APTES)和十八烷基三氯硅烷(OTS)在玻璃表面进行生长,考察反应时间、反应浓度和可水解基团对自组装单分子层膜的影响。用接触角仪和全反射红外光谱仪(FT-IR)对单分子膜进行表征。结果表明:带有—Cl水解基团的OTS最易生长,而带有乙氧基离去基团的OTES比带有甲氧基的DTMS容易反应。在24℃时,1 mmol/L OTES反应20 min自组装单分子层膜生长很好,并且表面比较规整均一。     

Rim instability in polymethylmethacrylate films on self-assembled monolayers with the hydrophilicity

The solvent-induced dewetting of polymethylmethacrylate (PMMA) films prepared on self-assembled monolayers (SAMs) supported by Si substrates was investigated with the hydrophilicity of the SAMs. Dewetting of the PMMA film was characterized as a function of the surface energy of the SAMs. At lower surface energies, the rim instability was found to be of type B. Type B instability created undulations in the rims of holes. As the holes merged, fingers and then droplets formed. At higher surface energies, the rim instability of the PMMA films was of type A. Type A instability did not form fingers. The formation of PMMA droplets was dominated by the total spreading coefficient (S) of the PMMA film on the SAMs, which can be expressed as the sum of the dispersion (S^d) and the polar components (S^p) of the spreading coefficient. S^d of the PMMA films decreased, whereas S^p increased as ultraviolet (UV) irradiation for periods of time between 0 s and 100 s, respectively. This result suggests that S^d destabilized but S^p stabilized the PMMA films.     

N-Hydroxysuccinimide-terminated self-assembled monolayers on gold for biomolecules immobilisation

Pure and mixed N-hydroxysuccinimide-terminated and butanethiol monolayers were prepared on flat gold (1 1 1) surfaces with the intent of developing suitable platforms for the direct biomolecules immobilisation. The self-assembled monolayers (SAMs) were characterised by electrochemical reductive desorption of the thiolate from the gold surface. The data have shown that certain solution proportions of the two compounds yield modified electrodes exhibiting intermediate electrochemical behaviour of the corresponding pure SAMs. The reactivity of the terminal N-hydroxysuccinimide (NHS) towards amine functionalities has been tested for the covalent attachment of Dopamine. The cyclic voltammetric responses of the investigated monolayers, after contacting with a Dopamine solution, have confirmed the chemical coupling of the amine as well as the formation of mixed SAMs. The Dopamine surface coverage increased with the amount of surface NHS. Laccase was also successfully immobilised onto this modified electrodes. The electrochemical behaviour of the modified SAMs with Laccase indicates direct electron transfer between the immobilised enzyme and the gold surface. Evidence for Laccase immobilisation was also provided by atomic force microscopic measurements.