A tripodal quinone-cyanine dye having one donor and three acceptors, that is, one quinone and three N-methylbenzothiazolium moieties, QCy­(MeBT)3, was synthesized by simple Knoevenagel condensation between 2-hydroxybenzene-1,3,5-tricarbaldehyde and N-methyl-2-methylbenzothiazolium iodide. The 700 nm (λex, 570 nm) and 600 nm (λex, 470 nm) fluorescence emission of QCy­(MeBT)3 was significantly and individually enhanced with the addition of G-quadruplex (G4) DNA and double-stranded DNA (dsDNA), respectively. The results of docking simulations and the response against the viscosity change revealed that the dual-fluorescence response was caused by the difference in the binding mode of QCy­(MeBT)3 depending on the DNA structure. The results of fluorescence microscopy imaging experiments using QCy­(MeBT)3 suggested that G4 DNAs and dsDNAs in the cell nucleus can be imaged with near-infrared (NIR, 700 nm) and red (600 nm) fluorescence emissions. Furthermore, pyridostatin-induced G4 formation in the living cells can be imaged with NIR fluorescence. The results indicated that QCy­(MeBT)3 has huge potential to be a NIR-fluorescent molecular probe for analyzing the structural dynamics of nucleic acids in living cells with a normal fluorescence microscope.

Bacteriorhodopsin (BR) functions as a light-driven proton pump that transitions between different states during the photocycle, such as all-trans (AT; BR568) and 13-cis, 15-syn (CS; BR548) state and K, L, M1, M2, N, and O intermediates. In this study, we used in situ photoirradiation 13C solid-state NMR to observe a variety of photo-intermediates and photoreaction pathways in [20-13C]retinal-WT-BR and its mutant [20-13C, 14-13C]retinal-D96N-BR. In WT-BR, the CS state converted to the CS* intermediate under photoirradiation with green light at −20 °C and consequently converted to the AT state in the dark. The AT state converted to the N intermediate under irradiation with green light. In D96N-BR, the CS state was converted to the CS* intermediate at −30 °C and consequently converted to the AT state. Simultaneously, the AT state converted to the M and L intermediates under green light illumination at −30 °C and subsequently converted to the AT state in the dark. The M intermediate was directly excited to the AT state by UV light illumination. We demonstrated that short-lived photo-intermediates could be observed in a stationary state using in situ photoirradiation solid-state NMR spectroscopy for WT-BR and D96N-BR, enabling insight into the light-driven proton pump activity of BR.