Investigating DNA-Mediated Charge Transport by Time-Resolved Spectroscopy

摘要: In all organisms, oxidation threatens the integrity of genome. Numerous studies have suggested that DNA-mediated charge transport (CT) may play an important role in sequestration, detection, and repair oxidative damage. To fully understand mechanism CT, it is necessary to characterize transient intermediates arise during reaction determine lifetimes these intermediates. Time-resolved spectroscopy most appropriate experimental method for such observations. Each intermediate has a characteristic spectrum. By observing time-dependent changes absorption spectrum sample, therefore possible what species are present at particular time how long exists solution. Experiments presented here involve use time-resolved better process CT. The study CT requires robust consistent triggering reaction. metal complexes traditionally been used this purpose provide several advantages over organic phototriggers: they synthetically versatile, stable solution, exhibit rich photophysics, many strong photooxidants. However, spectroscopic features follow photochemical processes triggered by probes generally broad optical bands. These can be difficult resolve samples contain absorbing species. For reason, we developed Re photooxidant bearing set vibrationally active carbonyl ligands covalently tethered DNA. Unlike bands visible range, vibrational narrow, well-resolved, specific. Such complex photophysical pathways observed biochemical systems with good precision, making them useful Specifically, [Re(CO)3(dppz)(py′-OR)]+ (dppz = dipyrido[3,2-a:2′,3′-c]-phenazine; py′-OR 4-functionalized pyridine) offers IR sensitivity oxidize DNA directly from excited state. behavior covalent noncovalent Re-DNA constructs was monitored (TRIR) UV/visible spectroscopies, as well methods, confirming ability trigger long-range Optical excitation leads population metal-to-ligand transfer states least two distinct intraligand states. Several observations injection Re*. include similarity between TRIR spectra reduced spectroelectrochemistry, appearance guanine radical signal spectra, eventual formation permanent products. majority reactivity occurs on ultrafast scale, although dependent slower conformational motions DNA, accumulation damage guanine, also observed. photooxidation activity compared other metallointercalators previously our laboratory [Rh(phi)2(bpy′)]3+ (phi 9,10-phenanthrenequinone diimine; bpy′ 4-methyl-4′-(butyric acid)-2,2′-bipyridine), [Ir(ppy)2(dppz′)]+ (ppy 2-phenylpyridine; dppz′ 6-(dipyrido[3,2-a:2′,3′-c]phenazin-11-yl)hex-5-ynoic acid), [Re(CO)3(dppz)(py′-OH)]+ (py′-OH 3-(pyridin-4-yl)-propanoic acid) were each Biochemical show upon irradiation, three efficiency: Rh > Ir. Comparison obtained spectroelectrochemistry after bulk reduction free those (TA) conjugates suggests 355 nm results Electrochemical experiments kinetic analysis TA decays verify primary factors responsible trend yield thermodynamic driving force variations efficiency back electron transfer, coupling redox-active DNA-binding proteins act hole sinks studied spectroscopy. designed support utilization biological systems. involving cell cycle regulator p53, photoexcitation weak band 405 nm. This band, which not lacking protein, resembles spectral feature tyrosine cation radical. Although reproducibility inconsistent, suggest photolysis sample p53. Similar conducted transcriptional activator SoxR. Here, presence dithionite, required solution keep protein reduced, complicates photochemistry system considerably. Regardless, absorbance 418 develops following provides evidence protein. base excision endonuclease III oxidants. flash-quench studies, addition negative 410 traces. direct Rh, Ir, complexes, no new transients detected but intensities resultant steady-state indicate taking place. described comprise developments story First, proof concept given valuable vibrationally-active probe. Further modifications characteristics further will allow us observe high resolution. Second, comparison probe established photooxidants shows its own right added growing toolbox phototriggers. Third, provided preliminary serve themselves. refinement methods greater sensitivity, increasing knowledge applications