Tianming Yao

A label-free fluorescent probe for Hg2+ and biothiols based on graphene oxide and Ru-complex

A novel, selective and sensitive switch-on fluorescent sensor for Hg2+ and switch-off fluorescent probe for biothiols was developed by using [Ru(bpy)2(pip)]2+ as the signal reporter and graphene oxide (GO) as the quencher. Due to the affinity of GO towards single-stranded DNA (ss-DNA) and [Ru(bpy)2(pip)]2+, the three components assembled, resulting in fluorescence quenching. Upon addition of Hg2+, a double-stranded DNA (ds-DNA) via T–Hg2+–T base pairs was formed, and [Ru(bpy)2(pip)]2+ intercalated into the newly formed ds-DNA. Then, [Ru(bpy)2(pip)]2+ and ds-DNA were removed from the surface of GO, resulting in the restoration of fluorescence. Subsequently, upon addition of biothiols, Hg2+ was released from ds-DNA, due to the higher affinity of Hg2+ to the sulfur atoms of biothiols, which could induce ds-DNA unwinding to form ss-DNA. Then ss-DNA and [Ru(bpy)2(pip)]2+ were adsorbed on the surface of GO, the fluorescence of [Ru(bpy)2(pip)]2+ was quenched again. Therefore, the changes in emission intensity of [Ru(bpy)2(pip)]2+ directly correlated to the amount of detection target (Hg2+ or biothiols) in solution. The assay exhibited high sensitivity and selectivity, with the limits of detection for Hg2+, cysteine (Cys) and glutathione (GSH) to be 2.34 nM, 6.20 nM and 4.60 nM, respectively.

Cu2+ modulated silver nanoclusters as an on–off–on fluorescence probe for the selective detection of l-histidine

In the present study, a new strategy based on Cu(2+) mediated DNA-templated silver nanoclusters (DNA-Ag NCs) was developed, as a label-free, on-off-on fluorescent probe for the detection of l-histidine. Eight synthesized DNA oligonucleotides (D1-D8) were experimentally tested, and D5-Ag NCs was finally selected for l-histidine detection due to its best fluorescent properties. The fluorescence emission of D5-Ag NCs could be quenched by Cu(2+) via electron or energy transfer. Upon addition of l-histidine, the chelation between Cu(2+) and the imidazole group of l-histidine leads to Cu(2+) liberation from D5-Ag NCs, and subsequently results in a dramatic fluorescence enhancement of the probe. The method displayed a good selectivity toward l-histidine over all the other amino acids, with a linear relationship in the range of 0.20-80μM, and a limit of detection (LOD) of 4.3nM. The strategy was also successfully applied to detect l-histidine in diluted human urine, exhibiting great opportunities for practical application in biological system.

A universal label-free fluorescent aptasensor based on Ru complex and quantum dots for adenosine, dopamine and 17β-estradiol detection

Based on specific aptamer binding properties, a strategy for adenosine, dopamine and 17β-estradiol detection was realised by employing Ru complex and quantum dots (QDs) as fluorescence probes. Ru complex, which could quench the fluorescence of QDs, preferred to bind with aptamer DNA and resulted in the fluorescence rise of QDs. When the aptamer DNA was incubated with the target first, it could not bind with Ru complex and the fluorescence of QDs was quenched. Under the optimal condition, the fluorescence intensity was linearly proportional to the concentration of adenosine, dopamine and 17β-estradiol with a limit of detection (LOD) of 101 nM, 19 nM and 37 nM, respectively. The experiments in fetal bovine serum were also carried out with good results. This universal method was rapid, label-free, low-cost, easy-operating and highly repeatable for the detection of adenosine, dopamine and 17β-estradiol. Qualitative detection by naked eyes was also available without complex instruments. It could also be extended to detect various analytes, such as metal ions, proteins and small molecules by using appropriate aptamers.

Graphene oxide–Ru complex for label-free assay of DNA sequence and potassium ions viafluorescence resonance energy transfer

Herein using classic DNA binding Ru polypyridine complex and graphene oxide, we developed a label-free and simple method for fluorescence recognition of complementary double-stranded DNA and aptamer-based potassium ions detection processes.

A comparative study of the interaction of two structurally analogous ruthenium complexes with human telomeric G-quadruplex DNA

Two new polypyridine ligands and their corresponding ruthenium(II) complexes have been prepared and characterized. The interactions of both complexes with human telomere quadruplex DNA (both the antiparallel basket and the mixed-hybrid G-quadruplex) have been studied by circular dichroism (CD), CD melting, UV-visible (UV-Vis), fluorescent intercalator displacement (FID) assays and molecular docking studies. The results show that both complexes can stabilize G-quadruplexes DNA and two complexes show different binding affinity for different G-quadruplexes DNA. The 1:1 stoichiometry was confirmed in the buffered solutions by the UV-Vis spectrophotometer using Jobs plot method and molecular docking studies. We have also investigated the interaction between the complexes and duplex DNA to gain some insight into the selectivity of the complexes for G-quadruplex structures. FID studies have shown that the complexes have a modest selectivity for G-quadruplex versus duplex DNA.

A molecular light switch Ru complex and quantum dots for the label-free, aptamer-based detection of thrombin

A simple, label-free method for the detection of thrombin has been developed based on the conformational transition of aptamer in the presence of the target by using a molecular light switch, Ru polypyridine complex, and quantum dots as novel fluorescence probes in aqueous solution.

Molecular Hairpin: A Possible Model for Inhibition of Tau Aggregation by Tannic Acid

Inhibition of anomalous aggregation of tau protein would be an attractive therapeutic target for Alzheimers disease (AD). In this study, tannic acid (TA), a polymeric plant polyphenol, and its monomer, gallic acid (GA), were introduced as the references to afford a molecular framework that integrates tau binding properties and inhibitory effects. Using a thioflavin S fluorescence assay and electron microscopy, we demonstrated that TA could competently inhibit the in vitro aggregation of tau peptide R3, corresponding to the third repeat unit of the microtubule-binding domain, with an IC50 of 3.5 μM, while GAs inhibition was comparatively piddling (with an IC50 of 92 μM). In the isothermal titration calorimetry experiment, we found that TA could strongly bind to R3 with a large amount of heat released. Circular dichroism spectra showed TA dose-dependently suppressed the conformational transition of R3 from a random coil structure to a β-sheet structure during the aggregation process. Finally, a structural model was built using molecular docking simulation to elucidate the possible binding sites for TA on the tau peptide surface. Our results suggest that TA recognizably interacts with tau peptide by forming a hairpin binding motif, a key framework required for inhibiting tau polymerization, in addition to hydrogen bonding, hydrophilic-hydrophobic interactions, and static electrical interactions, as reported previously. The inhibitory effect of TA on human full-length tau protein (tau441) was also verified by electron microscopy. This finding hints at the possibility of TA as a leading compound of anti-AD drugs and offers a new stratagem for the rational molecular design of a tau aggregation inhibitor.

Three label-free thrombin aptasensors based on aptamers and [Ru(bpy)2(o-mopip)]2+

Early disease diagnosis is very important for the prevention or mitigation of metastasis. Effective and efficient methods are needed to improve the diagnosis and assessment of diseases. Thrombin is a biomarker for diagnosis of some diseases, such as pulmonary metastases and diseases associated with coagulation abnormalities. Lots of methods for detecting thrombin have been reported, however, most of them were based on a single aptamer and needed modification of aptamers, and label free sensors for thrombin detection based on an aptamer pair are challenging. In this work, we present three label free sensing platforms based on [Ru(bpy)2(o-mopip)]2+ (bpy = 2,2-bipyridine; o-mopip = 2-(2-methoxylphenyl)imidazo[4,5-f][1,10]phenanthroline) (OMO) and graphene oxide (GO) for thrombin detection. Thrombin detection using our three label free sensors is accomplished by specific target recognition based on aptamer pair TBA1/TBA2-thrombin, single aptamer TBA1-thrombin or TBA2-thrombin, respectively. Direct readouts of the target recognition is achieved by restoration of the fluorescence of OMO prequenched by GO. All of the three sensing platforms exhibited high sensitivity and selectivity. Furthermore, all sensing platforms were successfully applied to thrombin analysis in diluted bovine serum. According to the thrombin dependent response of the three platforms, the sensing platform based on aptamer pair TBA1 and TBA2 showed the highest sensitivity, widest linearity, best selectivity and recovery in diluted serum to thrombin. These results show that the sensing platform based on an aptamer pair have great potential for clinical diagnosis of disease-related biomarkers.

Targeting human telomeric G-quadruplex DNA and inhibition of telomerase activity with [(dmb)2Ru(obip)Ru(dmb)2](4+).

Inhibition of telomerase by inducing/stabilizing G-quadruplex formation is a promising strategy to design new anticancer drugs. We synthesized and characterized a new dinuclear complex [(dmb)2Ru(obip)Ru(dmb)2]4+ (dmb = 4,4’-dimethyl-2,2’-bipyridine, obip = (2-(2-pyridyl)imidazo[4,5-f][1,10]phenanthroline) with high affinity for both antiparallel and mixed parallel / antiparallel G-quadruplex DNA. This complex can promote the formation and stabilize G-quadruplex DNA. Dialysis and TRAP experiments indicated that [(dmb)2Ru(obip)Ru(dmb)2]4+ acted as an excellent telomerase inhibitor due to its obvious selectivity for G-quadruplex DNA rather than double stranded DNA. In vitro co-culture experiments implied that [(dmb)2Ru(obip)Ru(dmb)2]4+ inhibited telomerase activity and hindered cancer cell proliferation without side effects to normal fibroblast cells. TUNEL assay indicated that inhibition of telomerase activity induced DNA cleavage further apoptosis in cancer cells. Therefore, RuII complex represents an exciting opportunity for anticancer drug design by specifically targeting cancer cell G-quadruplexes DNA.

Mercury(II) promotes the in vitro aggregation of tau fragment corresponding to the second repeat of microtubule‐binding domain: Coordination and conformational transition

The loss of metal homeostasis and the toxic effect of metal ion are important events in neurodegenerative and age-related diseases, such as Alzheimers disease (AD). For the first time, we investigated the impacts of mercury(II) ions on the folding and aggregation of Alzheimers tau fragment R2 (residues 275-305: VQIIN KKLDL SNVQS KCGSK DNIKH VPGGGS), corresponding to the second repeat unit of the microtubule-binding domain, which was believed to be pivotal to the biochemical properties of full tau protein. By ThS fluorescence assay and electron microscopy, we found that mercury(II) dramatically promoted heparin-induced aggregation of R2 at an optimum molar ratio of 1: 2 (metal: protein), and the resulting R2 filaments became smaller. Isothermal titration calorimetry (ITC) experiment revealed that the strong coordination of mercury(II) with R2 was an enthalpy-controlled, entropy-decreased thermodynamic process. The exceptionally large magnitude of heat release (ΔH₁ = -34.8 Kcal mol⁻¹) suggested that the most possible coordinating site on the R2 peptide chain was the thiol group of cysteine residue (Cys291), and this was further confirmed by a control experiment using Cys291 mutated R2. Circular dichroism spectrum demonstrated that this peptide underwent a significant conformational change from random coil to β-turn structure upon its binding to mercury(II) ion. This study was undertaken to better understand the mechanism of tau aggregation, and evaluate the possible role of mercury(II) in the pathogenesis of AD.