研究目的
To develop a controllable axial coordination system based on G-quadruplex/hemin complex for imaging intracellular nitric oxide and monitoring its relationship with p53 protein activity in living cells.
研究成果
The single-sided axial coordination system of G4/hemin complex was successfully constructed as a molecular switch for in situ imaging and detection of intracellular nitric oxide. The system demonstrated more powerful potential in aqueous axial ligation compared to the uncomplexed hemin, providing a new opportunity to understand the complicated physiological pathway of NO.
研究不足
The study is limited by the complexity of controlling single or dual axial interaction between the metal center of metalloporphyrin and the ligand, which usually requires complicated chemical modifications of the metalloporphyrin.
1：Experimental Design and Method Selection:
The study involved designing a series of fluorescent derivatives to interact with G-quadruplex/hemin complex via axial coordination. The mechanism of axial coordination was investigated using fluorescence quenching and recovery upon addition of nitric oxide.
2：Sample Selection and Data Sources:
The G4/hemin complex was prepared by incubating a parallel G-quadruplex with hemin. Fluorescent ligands were synthesized by conjugating fluorescein or coumarin fluorophore with amino-contained pyridine/imidazole derivatives.
3：List of Experimental Equipment and Materials:
The study utilized UV/vis absorption spectroscopy, fluorescence spectroscopy, mass spectrometry, X-ray photoelectron spectroscopy, electron spin resonance spectrum, and resonance Raman spectrum for characterization.
4：Experimental Procedures and Operational Workflow:
The G4/hemin complex was characterized with circular dichroism spectra. The axial coordination effect was investigated by synthesizing five different ligands and observing their fluorescence quenching efficiency. The competitive axial coordination with nitric oxide was studied to achieve fluorescence recovery.
5：Data Analysis Methods:
The binding constant was determined by UV-vis spectra titration. The structure of G4/hemin?NO complex was investigated using X-ray photoelectron spectroscopy, electron spin resonance spectrum, and resonance Raman spectrum.
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