Abstract
Vibrational spectroscopy is uniquely able to characterize protein dynamics and microenvironmental heterogeneity because it possesses an inherently high temporal resolution and employs probes of ultimately high structural resolution—the bonds themselves. The use of carbon–deuterium (C–D) bonds as vibrational labels circumvents the spectral congestion that otherwise precludes the use of vibrational spectroscopy to proteins and makes the observation of single vibrations within a protein possible while being wholly non-perturbative. Thus, C–D probes can be used to site-specifically characterize conformational heterogeneity and thermodynamic stability. C–D probes are also uniquely useful in characterizing the electrostatic microenvironment experienced by a specific residue side chain or backbone due to its effect on the C–D absorption frequency. In this chapter we describe the experimental procedures required to use C–D bonds and FT IR spectroscopy to characterize protein dynamics, structural and electrostatic heterogeneity, ligand binding, and folding.
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Acknowledgment
This work was supported by the National Science Foundation under Grant No. 0346967.
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Zimmermann, J., Romesberg, F.E. (2014). Carbon–Deuterium Bonds as Non-perturbative Infrared Probes of Protein Dynamics, Electrostatics, Heterogeneity, and Folding. In: Livesay, D. (eds) Protein Dynamics. Methods in Molecular Biology, vol 1084. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-658-0_6
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DOI: https://doi.org/10.1007/978-1-62703-658-0_6
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