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Förster Resonance Energy Transfer and Conformational Stability of Proteins. An Advanced Biophysical Module for Physical Chemistry Students
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Abstract
Protein folding is an exploding area of research in biophysics and physical chemistry. Here, we describe the integration of several techniques, including absorption spectroscopy, fluorescence spectroscopy, and Förster resonance energy transfer (FRET) measurements, to probe important topics in protein folding. Cytochrome c is used as a model protein; comparison of conformational stabilities (ΔGH2O) measured via two chemical denaturants, urea and guanidinium hydrochloride, illustrate important concepts in protein folding and intermolecular interactions. In addition, the determination of intraprotein distances based upon the FRET pair Trp-59 and the heme group for unfolded states of cytochrome c highlights the evolution of the protein structure under unfolding conditions. Analysis and discussion of these results provide opportunities to gain in-depth understanding of models for protein folding while enhancing students' skills with optical techniques. Collectively, the combination of optical spectroscopy, rigorous quantitative analysis, and a focus on biophysics illustrates the significance of fundamental research at the growing intersection of chemistry, biology, and physics.
Keywords (Audience):
Upper-Division UndergraduateKeywords (Domain):
BiochemistryKeywords (Pedagogy):
Hands-On Learning / ManipulativesKeywords (Subject):
Biophysical ChemistryCiting Articles
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This article has been cited by 3 ACS Journal articles (3 most recent appear below).
Femtosecond UV Studies of the Electronic Relaxation Processes in Cytochrome c
Olivier Bräm, Cristina Consani, Andrea Cannizzo, and Majed CherguiThe Journal of Physical Chemistry B2011 115 (46), 13723-13730Femtosecond UV Studies of the Electronic Relaxation Processes in Cytochrome c
Olivier Bräm, Cristina Consani, Andrea Cannizzo, and Majed CherguiThe Journal of Physical Chemistry B2011 115 (46), 13723-13730We report on an experimental study with UV and visible ultrafast time-gated emission and transient absorption of the early photodynamics of horse heart Cytochrome c in both ferric and ferrous redox states. A clear separation in time and energy of ...
Fluorescence Quenching, Lifetimes, and Fluorophore Solvent Accessibility
Lorenzo StellaJournal of Chemical Education2011 Article ASAPFluorescence Quenching, Lifetimes, and Fluorophore Solvent Accessibility
Lorenzo StellaJournal of Chemical Education2011 Article ASAPThe influence of the excited-state lifetime of a fluorophore on its susceptibility to collisional quenching is discussed. In the case of cytochrome c, FRET to the heme moiety strongly reduces the tryptophan fluorescence lifetime. This should be taken into ...
Quenching of Tryptophan Fluorescence in Unfolded Cytochrome c: A Biophysics Experiment for Physical Chemistry Students
Diana E. Schlamadinger, Dina I. Kats and Judy E. KimJournal of Chemical Education2010 87 (9), 961-964Quenching of Tryptophan Fluorescence in Unfolded Cytochrome c: A Biophysics Experiment for Physical Chemistry Students
Diana E. Schlamadinger, Dina I. Kats and Judy E. KimJournal of Chemical Education2010 87 (9), 961-964Laboratory experiments that focus on protein folding provide excellent opportunities for undergraduate students to learn important topics in the expanding interdisciplinary field of biophysics. Here, we describe the use of Stern−Volmer plots to determine ...
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- Received: August 03, 2009
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