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Distance-dependent fluorescence quenching ofN-acetyl-L-tryptophanamide by acrylamide
B Zelent1, J Kuśba, I Gryczynski
1Center for Fluorescence Spectroscopy, Department of Biological Chemistry, University of Maryland at Baltimore, School of Medicine, 108 North Greene Street, 21201, Baltimore, Maryland.
Journal of Fluorescence
|November 16, 2013
Summary
Acrylamide quenching of N-acetyl-L-tryptophanamide (NATA) is distance-dependent, not diffusion-limited. This through-space interaction explains observed data and impacts protein quenching studies.
Area of Science:
- Photochemistry
- Biophysical Chemistry
- Spectroscopy
Background:
- N-acetyl-L-tryptophanamide (NATA) is a model fluorophore.
- Acrylamide is a common quencher used in fluorescence studies.
- Understanding quenching mechanisms is crucial for interpreting biomolecular interactions.
Purpose of the Study:
- To investigate the quenching mechanism of NATA by acrylamide in propylene glycol at various temperatures.
- To determine if the quenching process is diffusion-limited or follows other models.
- To elucidate the role of distance-dependent interactions in fluorescence quenching.
Main Methods:
- Time-dependent intensity decay measurements of NATA.
- Frequency-domain and steady-state fluorescence spectroscopy.
- Analysis using Stern-Volmer plots and comparison with the Collins-Kimball model.
Main Results:
- NATA intensity decays became heterogeneous with acrylamide presence.
- Observed quenching in vitrified propylene glycol, indicating non-diffusional quenching.
- Stern-Volmer plots showed upward curvature, inconsistent with simple diffusion models.
Conclusions:
- Quenching is primarily a through-space interaction, with a rate constant exponentially dependent on distance.
- The Collins-Kimball model is inadequate for describing this quenching.
- Distance-dependent quenching rates are important for interpreting acrylamide quenching of proteins.
