title: Scanning Single-Molecule Fluorescence Correlation Spectroscopy Enables Kinetics Study of DNA Hairpin Folding with a Time Window from Microseconds to Seconds.
abstract: Single-molecule fluorescence measurements have been widely used to explore kinetics and dynamics of biological systems. Among them, single-molecule imaging (SMI) is good at tracking processes slower than tens of milliseconds, whereas fluorescence correlation spectroscopy (FCS) is good at probing processes faster than submilliseconds. However, there is still shortage of simple yet effective single-molecule fluorescence method to cover the time-scale between submilliseconds and tens of milliseconds. To effectively bridge this millisecond gap, we developed a single-molecule fluorescence correlation spectroscopy (smFCS) method that works on surface-immobilized single molecules through surface scanning. We validated it by monitoring the classical DNA hairpin folding process. With a wide time window from microseconds to seconds, the experimental data are well fitted to the two-state folding model. All relevant molecular parameters, including the relative fluorescence brightness, equilibrium constant, and reaction rate constants, were uniquely determined.
authors.affiliation: Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Department of Chemical Biology, College of Chemistry and Molecular Engineering, and Biodynamic Optical Imaging Center (BIOPIC), Peking University , Beijing 100871, China.
authors.forename: Huimin
authors.initials: H
authors.lastname: Bi
authors.affiliation: Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Department of Chemical Biology, College of Chemistry and Molecular Engineering, and Biodynamic Optical Imaging Center (BIOPIC), Peking University , Beijing 100871, China.
authors.forename: Yandong
authors.initials: Y
authors.lastname: Yin
authors.affiliation: Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Department of Chemical Biology, College of Chemistry and Molecular Engineering, and Biodynamic Optical Imaging Center (BIOPIC), Peking University , Beijing 100871, China.
authors.forename: Bailong
authors.initials: B
authors.lastname: Pan
authors.affiliation: Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Department of Chemical Biology, College of Chemistry and Molecular Engineering, and Biodynamic Optical Imaging Center (BIOPIC), Peking University , Beijing 100871, China.
authors.forename: Geng
authors.initials: G
authors.lastname: Li
authors.affiliation: Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Department of Chemical Biology, College of Chemistry and Molecular Engineering, and Biodynamic Optical Imaging Center (BIOPIC), Peking University , Beijing 100871, China.
authors.forename: Xin Sheng
authors.initials: XS
authors.lastname: Zhao
mesh_terms: D004247:DNA / Q000737:chemistry*; D007700:Kinetics; D000072760:Single Molecule Imaging; D013050:Spectrometry, Fluorescence; D013997:Time Factors
chemical_list: D004247:DNA