Dynamics of biological macromolecules investigated by fluorescence depolarization
Piston, David William
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https://hdl.handle.net/2142/23944
Description
Title
Dynamics of biological macromolecules investigated by fluorescence depolarization
Author(s)
Piston, David William
Issue Date
1989
Doctoral Committee Chair(s)
Gratton, E.
Department of Study
Physics
Discipline
Physics
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
dynamics of biomolecular systems
biological macromolecules
fluorescence depolarization
time-resolved fluorescence
nanosecond dynamics
Language
en
Abstract
"The fast dynamics of biomolecular systems are believed to be important to their physiological function.
Fluorescence depolarization is a powerful technique for the investigation of these dynamics.
A general overview of the use of time-resolved fluorescence to investigate nanosecond dynamics is
presented. The basic theory and experimental techniques for measuring time-resolved fluorescence
properties in the frequency domain are outlined. Data obtained in fluorescence depolarization experiments
is highly complex. Mathematical models for analyzing data from depolarization due to
rotational motion have been largely based on the diffusion equation. It has been implicitly stated
that a ""jump"" model should give the same result for the anisotropy decay as the diffusion equation.
In this work, we derive the general result from the jump model, where the excitation and emission
dipoles are not necessarily coincident with any of the principal rotational axes of the fluorophore is
derived. This result is found to be different from that of the diffusion equation. This difference is
significant since, for systems where the fluorophore is not much larger than the solvent molecules
or where the molecule may be limited to a few preferred orientations (for example, residues in proteins),
the actual physical mechanism of rotation may not be accurately represented by continuous
diffusion. Since there are cases where symmetry causes the two models to agree, it is proposed that
both models are only limiting cases of the underlying physical process of rotational motion. The
physical assumptions behind the two models and the limits of applicability of each approach are
discussed, and some of the thermodynamic properties are considered. Finally, several applications
of the compartmental jump approach are presented: a free disk-like molecule, a rod-like molecule
in a phospholipid bilayer, and a tryptophan residue in a protein matrix."
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