Sensitive Detection of Polynucleotide Kinase Activity by Paper-Based Fluorescence Assay with λ Exonuclease Assistance

The phosphorylation of nucleic acid with 5′-OH termini catalyzed by polynucleotide kinase (PNK) involves several significant cellular events. Here a paper-based fluorescence assay with λ exonuclease assistance was reported for facile detection of PNK activity through monitoring the change of fluorescence intensity on paper surface. Cy5-labeled ssDNA was first immobilized on the surface of aldehyde group modified paper, and BHQ-labeled ssDNA was then employed to quench the fluorescence of the immobilized Cy5-labeled ssDNA with the help of an adaptor ssDNA. When PNK and λ exonuclease cleavage reaction were introduced, the fluorescence quenching effect on the paper surface was blocked because of the digestion of phosphorylated dsDNA by the coupled enzymes. By using this paper-based assay, PNK activity both in pure reaction buffer and in practical biosample have been successfully measured. Highly sensitive detection of PNK activity down to 0.0001 U mL^–1 and lysate of about 50 cells is achieved. The inhibition of PNK activity has also been investigated and a satisfactory result is obtained

Geminal Tetraauration of Acetonitrile: Hemilabile-Phosphine-Stabilized Au₈Ag₄ Cluster Compounds

Unprecedented geminal tetraauration of acetonitrile has been realized through C–H activation by Au­(I)–Ag­(I) clusters under mild conditions. The reaction of [OAu₃Ag­(dppy)₃]­(BF₄)₂ (dppy = diphenylphosphino-2-pyridine) (<b>1</b>), AgBF₄, and acetonitrile in the presence of methanol at room temperature resulted in the isolation of the novel cluster [(CCN)₂Au₈Ag₄(dppy)₈(CH₃CN)₂]­(BF₄)₆ (<b>2</b>). The centrosymmetric structure consists of two Au₄Ag₂ motifs stabilized by hemilabile phosphines. Triply deprotonated acetonitrile (CCN^3–) is found in a Au₄Ag environment with the terminal carbon bridging four Au­(I) centers and the nitrogen donor linking a Ag­(I) ion, which is the first example of a μ₅-CCN^3– coordination mode. A concerted metalation/deprotonation process for the C–H activation of acetonitrile that indicates the importance of the oxo ion of the oxonium Au­(I) cluster is proposed. Cluster <b>2</b> emits bright green light in the solid state at room temperature upon UV irradiation

Gelling properties of myosin as affected by L-lysine and L-arginine by changing the main molecular forces and microstructure

<p>This paper investigated the effects of l-lysine (Lys) and l-arginine (Arg) on the water-holding capacity (WHC) and hardness of myosin gel as well as the mechanism of these effects. The results showed that Lys, Arg, and KOH increased the WHC but decreased the hardness in all cases. At pH 6.32, the WHC increased in the order control < KOH < Lys ≈ Arg, while the hardness decreased in the order control > KOH > Lys > Arg. Lys shortened low field nuclear magnetic resonance spin-spin relaxation times (T₂), while Arg and KOH slightly affected T₂. Lys, Arg, and KOH had different influences on the molecular forces in myosin gels and the distribution of myosin size. Lys formed a gel with porous clusters, while Arg generated a gel with fine caves. Therefore, both Lys and Arg changed the microstructure of myosin gel by changing the distribution of the myosin size as well as the molecular forces that form and/or maintain myosin gel, ultimately contributing to the differences in the WHC, hardness, and T₂.</p

Low-Potential Synthesis of “Clean” Au Nanodendrites and Their High Performance toward Ethanol Oxidation

The shape control of Au nanocrystals is crucial to their catalytic applications and optical properties. Well-defined Au nanodendrites (NDs) have been prepared on a glassy carbon electrode using low-potential synthesis, assisted by ethylenediamine (EDA). The effects of applied potential, deposition time, and HAuCl₄ (or EDA) concentrations on the morphology of the Au deposits are discussed in our work. The growth mechanism can be explained by a two-staged growth of dendrites: initial branching and subsequent dendritic growth. The Au NDs exhibits superior catalytic performance toward ethanol oxidation, in comparison with the polycrystalline Au nanoparticles. The simple and facile synthetic technique can be applied to the construction of other metals with complex hierarchical structures on a large-scale

Geminal Tetraauration of Acetonitrile: Hemilabile-Phosphine-Stabilized Au₈Ag₄ Cluster Compounds

Unprecedented geminal tetraauration of acetonitrile has been realized through C–H activation by Au­(I)–Ag­(I) clusters under mild conditions. The reaction of [OAu₃Ag­(dppy)₃]­(BF₄)₂ (dppy = diphenylphosphino-2-pyridine) (<b>1</b>), AgBF₄, and acetonitrile in the presence of methanol at room temperature resulted in the isolation of the novel cluster [(CCN)₂Au₈Ag₄(dppy)₈(CH₃CN)₂]­(BF₄)₆ (<b>2</b>). The centrosymmetric structure consists of two Au₄Ag₂ motifs stabilized by hemilabile phosphines. Triply deprotonated acetonitrile (CCN^3–) is found in a Au₄Ag environment with the terminal carbon bridging four Au­(I) centers and the nitrogen donor linking a Ag­(I) ion, which is the first example of a μ₅-CCN^3– coordination mode. A concerted metalation/deprotonation process for the C–H activation of acetonitrile that indicates the importance of the oxo ion of the oxonium Au­(I) cluster is proposed. Cluster <b>2</b> emits bright green light in the solid state at room temperature upon UV irradiation

Poly(glycidyl methacrylate-<i>co</i>-2-hydroxyethyl methacrylate) Brushes as Peptide/Protein Microarray Substrate for Improving Protein Binding and Functionality

We developed a three-dimensional (3D) polymer-brush substrate for protein and peptide microarray fabrication, and this substrate was facilely prepared by copolymerization of glycidyl methacrylate (GMA) and 2-hydroxyethyl methacrylate (HEMA) monomers via surface-initiated atom transfer radical polymerization (SI-ATRP) on a glass slide. The performance of obtained poly­(glycidyl methacrylate-<i>co</i>-2-hydroxyethyl methacrylate) (P­(GMA-HEMA)) brush substrate was assessed by binding of human IgG with rabbit antihuman IgG antibodies on a protein microarray and by the determination of matrix metalloproteinase (MMP) activities on a peptide microarray. The P­(GMA-HEMA) brush substrate exhibited higher immobilization capacities for proteins and peptides than those of a two-dimensional (2D) planar epoxy slide. Furthermore, the sensitivity of the P­(GMA-HEMA) brush-based microarray on rabbit antihuman IgG antibody detection was much higher than that of its 2D counterpart. The enzyme activities of MMPs were determined specifically with a low detection limit of 6.0 pg mL^–1 for MMP-2 and 5.7 pg mL^–1 for MMP-9. By taking advantage of the biocompatibility of PHEMA, the P­(GMA-HEMA) brush-based peptide microarray was also employed to evaluate the secretion of MMP-2 and MMP-9 by cells cultured off the chip or directly on the chip, and satisfactory results were obtained

Modular Construction/Off-Site Construction

Modular Construction/Off-Site Constructio

Solvent Dependent Excited State Behaviors of Luminescent Gold(I)–Silver(I) Cluster with Hypercoordinated Carbon

Polynuclear Au­(I) complexes continues to attract considerable attention because of their bright emissions in the visible wavelength, which hold promise in applications in luminescence, fluorescence sensing, and bioimaging. Despite various spectroscopic investigations on their steady state properties, detailed understanding of the origin of their emissions and excited state relaxations is still lacking. Here, we report femtosecond time-resolved transient absorption experiments combined with quantum chemical calculations on a brightly emissive [Au₆Ag₂(C)­(dppy)₆]­(BF₄)₄ cluster in different solvents. Global analysis on the transient absorption spectra based on a sequential model gives three spectral components: (1) excited state absorption (ESA) of ¹MLCT_Au state (τ = 1–3 ps); (2) ESA of ³MLCT_Au state (τ = 11–40 ps), and (3) ESA of ³MLCT_Ag state (long-lived). By variation of the solvent’s polarity and hydrogen bonding ability, the relative population of the triplet MLCT states and the emission properties can be modulated. Especially in methanol, an additional site specific O–H···π bond is formed between methanol molecules and aromatic rings of ligands, which enhances the ultrafast nonradiative decay from the hydrogen bond stabilized ³MLCT_Au state and reduces the population of the emissive ³MLCT_Ag state. The results presented here about the excited state dynamics of luminescent gold­(I)–silver­(I) cluster allow a deeper insight into the origin of their emissions by monitoring the population of the emissive ³MLCT_Ag state and dark ³MLCT_Au state in different environments

Evaluation of Matrix Metalloproteinase Inhibition by Peptide Microarray-Based Fluorescence Assay on Polymer Brush Substrate and in Vivo Assessment

Matrix metalloproteinases (MMPs) are important biomarkers and potential therapeutic targets of tumor. In this report, a peptide microarray-based fluorescence assay is developed for MMPs inhibitors evaluation through immobilization of biotin-modified peptides on the poly­(glycidyl methacrylate-<i>co</i>-2-hydroxyethyl methacrylate) (P­(GMA-HEMA)) brush-modified glass slides. After biotin is recognized with cyanine 3 (Cy3)-modified avidin (Cy3-avidin), the microarrays can produce strong fluorescence signal. The biotin moieties detach from microarray, when the biotin-modified peptide substrates are specially cleaved by a MMP, resulting in decreased fluorescence intensity of the microarray. The decreasing level of fluorescence intensity is correlated with the MMP inhibition. Nine known MMP inhibitors against MMP-2 and MMP-9 are evaluated by the assay, and the quantitative determination of inhibitory potencies (half maximal inhibitory concentration) are obtained, which are comparable with the literatures. Two biocompatible fluorogenic peptides containing MMP-specific recognition sequences and FAM/Dabcyl fluorophore-quencher pair are designed as activatable reporter probes for sensing MMP-2 and MMP-9 activities in cell and in vivo. The peptide microarray-based results are well verified by the cell inhibition assay and in vitro fluorescence imaging, and further confirmed by the in vivo imaging of HT-1080 tumor-bearing mice

Activation of GABA_A receptors attenuated the peak current amplitude and enhanced the sustained current of ASIC1a.

<p>A, Example traces of a fast-inactivating transient current and a sustained current of ASIC1a activated by pH 3.5. GABA (100 μM) attenuated a fast-inactivating transient current and enhanced the sustained current of ASIC1a in HEK293 cells co-transfected with GABA_A receptor subunits (α₁ and β₂) and ASIC1a, which can totally abolished by co-application of picrotoxin (100 μM) with GABA. ASIC1a current traces were superimposed to the right (inset) (B). C, GABA had no effect on ASIC1a currents in HEK293 cells transfected with ASIC1a cDNA only. ASIC1a current traces were superimposed to the right (inset).</p