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Switching of G-quadruplex (G4) structures between variant types of folding has been proved to be a versatile tool for regulation of genomic expression and development of nucleic acid-based constructs. Various specific ligands have been developed to target G4s in K+ solution with therapeutic prospects. Although G4 structures have been reported to be converted by sequence modification or a unimolecular ligand binding event in K+-deficient conditions, switching G4s towards non-G4 folding continues to be a great challenge due to the stability of G4 in physiological K+ conditions. Herein, we first observed the G4 switching towards parallel-stranded duplex (psDNA) by multimolecular ligand binding (namely ligand clustering) to overcome the switching barrier in K+. Purine-rich sequences (e.g. those from the KRAS promoter region) can be converted from G4 structures to dimeric psDNAs using molecular rotors (e.g. thioflavin T and thiazole orange) as initiators. The formed psDNAs provided multiple binding sites for molecular rotor clustering to favor subsequent structures with stability higher than the corresponding G4 folding. Our finding provides a clue to designing ligands with the competency of molecular rotor clustering to implement an efficient G4 switching.

DNA foldings provide variant possibilities to develop DNAzymes with remarkable catalytic performance. In spite of fruitful reports on G-quadruplex DNAzymes, four-stranded cytosine-rich i-motifs have not been explored as the potential skeletons of DNAzymes. In this work, we developed a visible light-driven DNAzyme based on human telomeric i-motifs using a natural photosensitizer of hypericin (Hyp) as the cofactor and dissolved oxygen as the oxidant source. The i-motif folding in acidic solution caused the distal thymine overhangs at the 3' and 5' ends to approach each other to provide a favorable binding site for Hyp via an interaction of fully complementary hydrogen bonding. However, the i-motifs without the distal overhangs or with the inappropriate overhang length and the base identity exhibited no binding with Hyp. The binding event converted Hyp from the fully dark state to the emissive state under visible light illumination. Subsequently, the excited Hyp had an opportunity to transfer energy to dissolved oxygen. Resultantly, singlet oxygen (

This study aimed to explore the interaction between bombykol and BmOR1 and also provide a paradigm for agroforestry pest control. The electrochemical biosensor signal amplification system was used: nanogold with horseradish peroxidase. An electrochemical bilayer nanogold membrane receptor sensor was developed using the following schemes and processes: twice self-assembly of nanogold and succeeding absorption of Bombyx mori olfactory receptor 1 (BmOR1); sex pheromone–binding protein; spectral scanning and transmission electron microscope to characterize nanogold sol; and atomic force microscope, cyclic voltammetry, and AC impedance methods to characterize individual processes of sensor assembly. The amperometric I–T curve was adopted to measure the response current upon interaction with different concentrations of bombykol (diluted in phosphate-buffered saline) and BmOR1. The results demonstrated the receptor–ligand interaction pattern, which was similar to enzymatic reaction kinetics, with the activation constant Ka of up to 8.57 × 10−20 mol/L and signal magnification of about 10,000-fold. In this study, the simulation of intracellular receptor signaling cascade by an electrochemical signal amplification system helped in directly measuring BmOR1–bombykol ligand interaction and exploring the kinetics after the self-assembly of BmOR1 on the biosensor. It provided a novel platform for future studies on receptor–ligand interaction.

Sweet and umami are respectively elicited by sweet/umami receptor on the tongue and palate epithelium. However, the molecular machinery allowing to taste reaction remains incompletely understood. Through a phosphoproteomic approach, we found the key proteins that trigger taste mechanisms based on the phosphorylation cascades. Thereinto, ryanodine receptor isoform 1 (RYR1) was further verified by sensor and behaviors assay. A model proposing RYR1-mediated sweet/umami signaling: RYR1 channel which mediates Ca2+ release from the endoplasmic reticulum is closed by its dephosphorylation in the bud tissue after umami/sweet treatment. And the alteration of Ca2+ content in the cytosol induces a transient membrane depolarization and generates cell current for taste signaling transduction. We demonstrate that RYR1 is a new channel in regulation of sweet/umami signaling transduction and also propose a “metabolic clock” notion based on sweet/umami sensing. Our study provides a rich fundamental for a system-level understanding of taste perception mechanism.

Trinucleotide repeats (TRs) with abnormal lengths and atypical folding are implicated in various neurodegenerative diseases. The least stable cytosine-cytosine (C–C) mismatches in TRs when structuring into homoduplexes/hairpins have more chance in certain sequence contexts to preferentially adopt an extrahelical (E-motif) conformation with respect to those in polarity-inverted intrahelical counterparts. Herein, we designed a trihydroxyphenyl porphyrin ligand (POH3) to meet the challenge towards resolving the E-motif conformation. POH3 exhibited a specific 2:1 binding with DNAs adopting the E-motif cytosine conformation, independent of the TRs length. The trihydroxyl pattern was very crucial to gain the E-motif selectivity over the polarity-inverted counterparts via the complementary hydrogen bonding that occurred in the minor groove. Our work first elucidates the rationale in designing ligands to selectively resolve the E-motif nucleotides within TRs.

Trihydroxyphenyl porphyrin (POH3) was designed to specifically bind with a triplex DNA resulting in a turn-on fluorescence response. This ensemble can be developed into a catalytic triplex DNAzyme towards porphyrin metalation. The catalytic activity is initiated by the enhanced basicity of POH3 upon binding with the triplex DNA.

GPR120 (FFAR4) is a free fatty acid receptor, which transmits a signal of excess fatty acid. When a person ingests too much fatty acid, free fatty acid appears in the intestine. At this time, it can sense the free fatty acid and thus control a person's appetite. In this study, we constructed three sensors for the detection of 14 natural fatty acids. Then the interconnected allosteric interactions (Ka) of the three sensors are compared, and their dynamics are also studied. The results show that all three receptor sensors can sense 14 fatty acids. And the Pearson correlation coefficient of Ka between the hGPR120 nanogold sensor and the hGPR120 self-assembled taste bud tissue sensor reached 0.966. This indicated that the hGPR120 nanogold sensor could mimick the ability of the hGPR120 receptor to transmit signals to cells and could be used to study intracellular signaling and activation of this receptor. We also found that the surface of the cattle taste bud tissue does’t contain the GPR120 receptor. We could use the differences between human and cattle taste bud tissues to develop self-assembled human receptor sensors to provide novel technologies and detection platforms for human receptor-ligand interaction studies and drug screening.

The G protein cascade amplification system couples with several receptors to sense/amplify the cellular signal, implying universal application. In order to explore whether GPCRs can trigger G protein signal amplification in tissues/cells from different species, bombykol receptor was isolated and purified from antennas of male Bombyx mori, which subsequently self-assembled on the cell membrane in rat taste buds/rat vomeronasa/catfish tentacles/taste bud tissues of rabbits/pig/cattle in those lacking endogenous bombykol receptor, followed by immobilization between two sheets of nucleopore membranes fixed by sodium alginate-starch gel, forming the sandwich-type sensing membrane, which in turn was immobilized on the glass-carbon electrode. Thus, bombykol receptor sensors were established with different tissues. The response current of bombykol receptor sensor toward bombykol was measured with an electrochemical workstation. Every bombykol receptor sensor could sense bombykol based on enzyme-substrate kinetics. The double reciprocal plot and the activation constant values of bombykol receptor sensors assembled with rat taste buds, rat vomeronasa, catfish tentacles, rabbit taste buds, pig taste buds, and cattle taste buds were calculated. Approximately 2-3 receptors could trigger the G protein cascade amplification system and achieve the maximum signal output. Moreover, the detection lower limit indicated that the bombykol receptor self-assembled on the cell membranes of different tissues that transmitted and amplified the bombykol signal with hypersensitivity. Also, cattle taste bud tissues served as an ideal system for heterogeneous GPCRs self-assembly and signal sensing/amplification. This sensing technique and method had promising potential in studies of biological pest control, sex pheromone detection, and receptor structure and function.

An electrochemical double-layer Au nanoparticle membrane immunosensor was developed using an electrochemical biosensing signal amplification system with Au nanoparticles, thionine, chitosan, and horseradish peroxidase, which was fabricated using double self-adsorption of Au nanoparticle sol followed by anti-α-fetoprotein Balb/c mouse monoclonal antibody adsorption. The AuNPs sol was characterized by spectrum scanning and transmission electron microscopy. The immunosensor was characterized by atomic force microscopy, cyclic voltammetry, and alternating-current impedance during each stage of adsorption and assembly. The amperometric I-t curve method was used to measure α-fetoprotein (AFP) diluted in phosphate buffered saline. The result indicated a wide linear range, and the change rate of steady-current before and after immune response had linear correlation within the range 0.1–104 pg/mL AFP. The current change rate equation was △I = 5.82334 lgC + 37.01195 (R2 = 0.9922). The lowest limit of detection was 0.03 pg/mL (S/N = 3), and the reproducibility of the sensor was good. Additionally, the sensor could be stably stored above phosphate buffered saline at 4 °C for more than 24 days. More importantly, the sensor is label-free, reagentless and low fouling, making it capable of assaying AFP in real serum samples without suffering from significant interference or biofouling.