Fluorescence Depolarization Study of Internal Tryptophan Mobility in Hydrated Azurin Films
Limkeman, Mark Kenneth
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https://hdl.handle.net/2142/77369
Description
Title
Fluorescence Depolarization Study of Internal Tryptophan Mobility in Hydrated Azurin Films
Author(s)
Limkeman, Mark Kenneth
Issue Date
1984
Department of Study
Physics
Discipline
Physics
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Physics, Molecular
Language
eng
Abstract
The effect of hydration on protein dynamics was investigated using steady state fluorescence depolarization on azurin, which has a single tryptophan residue well-buried in the hydrophobic interior. Azurin was imbedded in a thin, solid, water-permeable polymer film. This procedure inhibited whole-protein motion while allowing examination of internal degrees of rotational freedom in the protein as a function of film hydration. In the dry film at room temperature the tryptophan fluorescence emission showed the limiting polarization characteristic of low temperature solutions in which all motion is frozen. As the hydration was increased, the observed polarization value changed sharply at about 0.6 h (h = g water/g film), indicating an increase in the rotational mobility of the tryptophan. The fully hydrated film had a polarization equal to that of an azurin solution measurement extrapolated to infinite viscosity where the rotation of the protein as a whole is hindered. The conclusion is that the protein internal mobility is linked to the hydration level, and that hydration can turn on the full protein flexibility on the nanosecond time scale accessible by fluorescence techniques.
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