Fluorescent and phosphorescent probes that have readily interpretable emission properties can be specifically inserted into biological macromolecules to reveal facets of their structure and dynamics: (1) Proximity. Singlet-singlet and triplet-singlet energy transfer can serve as spectroscopic rulers in the 10-65 Å range, whereas triplet-triplet transfer can be used to show that two groups are less than about 12 Å apart. (2) Rotational mobility. Nanosecond fluorescence polarization measurements can reveal whether a macromolecular system has any modes of flexibility in times of nanoseconds. (3) Polarity. The presence of mobile dipoles in the environment of certain chromophores is reflected in their fluorescence quantum yield and emission spectrum
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