Orientation-dependent FRET system reveals differences in structures and flexibilities of nicked and gapped DNA duplexes

Abstract

Differences in structures and flexibilities of DNA duplexes play important roles on recognition by DNA-binding proteins. We herein describe a novel method for structural analyses of DNA duplexes by using orientation dependence of Förster resonance energy transfer (FRET). We first analyzed canonical B-form duplex and correct structural parameters were obtained. The experimental FRET efficiencies were in excellent agreement with values theoretically calculated by using determined parameters. We then investigated DNA duplexes with nick and gaps, which are key intermediates in DNA repair systems. Effects of gap size on structures and flexibilities were successfully revealed. Since our method is facile and sensitive, it could be widely used to analyze DNA structures containing damages and non-natural molecules.

Quantitative evaluation of energy migration between identical chromophores enabled by breaking symmetry

“…Generally, emission intensity increased as the number of base pairs increased; however, maxima of emission intensity were observed when 8 and 13 base pairs separated the fluorophores. Similar a 5-base-pair cycle of emission intensity was observed with hetero FRET from pyrene to perylene that corresponds to a half turn of the B-form DNA duplex 32,39 . Our result indicated that energy migration between identical chromophores also exhibits orientation dependence.…”

Quantitative Analyses of Förster Resonance Energy Transfer between Identical Pyrene Chromophores (Homo‐FRET) In DNA Scaffolds.

“…These duplexes were prepared by hybridizing three strands. We have reported that orientation dependence of hetero‐FRET kept intact in the case of nicked duplex [8b] . The number of base pairs between two pyrenes can be changed from 2 bp to 13 bp by hybridizing strands tethering dyes at different positions (Table S1).…”

Sulfonated Pyrene as a Photoregulator for Single‐Stranded DNA Looping

“…The formation of Py excimer is exceptionally sensitive to the surrounding microenvironment, making Py a gold probe of microenvironmental change . Hence, and owing to its chemical stability and charge‐carrier mobility, Py and its derivatives (Pys) have been widely used for studying the assembly, conformation, and dynamics of macromolecules and supramolecular assemblies of varied size, composition, and architecture . In most cases, techniques involving the processes of fluorescence resonance energy transfer (FRET) or changes in fluorescence (intensity and/or lifetime) from the singlet excited state of monomer ( 1 Py * ) and excimer ( 1 Py 2 * ) were employed .…”

“…Hence, and owing to its chemical stability and charge‐carrier mobility, Py and its derivatives (Pys) have been widely used for studying the assembly, conformation, and dynamics of macromolecules and supramolecular assemblies of varied size, composition, and architecture . In most cases, techniques involving the processes of fluorescence resonance energy transfer (FRET) or changes in fluorescence (intensity and/or lifetime) from the singlet excited state of monomer ( 1 Py * ) and excimer ( 1 Py 2 * ) were employed . The above‐mentioned techniques are dependent on the photophysical properties of 1 Py * and 1 Py 2 * , the lifetimes of which are reported to be around 400 ns (680 ns was also reported) and 60 ns (90 ns was also reported) in deoxygenated cyclohexane, respectively .…”