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Approaches for the measurement of solvent exposure in proteins by 19F NMR

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Abstract

Fluorine NMR is a useful tool to probe protein folding, conformation and local topology owing to the sensitivity of the chemical shift to the local electrostatic environment. As an example we make use of 19F NMR and 3-fluorotyrosine to evaluate the conformation and topology of the tyrosine residues (Tyr-99 and Tyr-138) within the EF-hand motif of the C-terminal domain of calmodulin (CaM) in both the calcium-loaded and calcium-free states. We critically compare approaches to assess topology and solvent exposure via solvent isotope shifts, 19F spin–lattice relaxation rates, 1H–19F nuclear Overhauser effects, and paramagnetic shifts and relaxation rates from dissolved oxygen. Both the solvent isotope shifts and paramagnetic shifts from dissolved oxygen sensitively reflect solvent exposed surface areas.

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Abbreviations

1D:

One-dimensional

CaM:

Calmodulin

CSA:

Chemical shift anisotropy

DNase:

Deoxyribonuclease

RNase:

Ribonuclease

ID:

Inner diameter

IPTG:

Isopropyl β-d-1-thiogalactopyranoside

EDTA:

Ethylenediaminetetraacetic acid

NMR:

Nuclear magnetic resonance

OD:

Outer diameter

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Acknowledgments

We wish to thank Professor Mitsu Ikura (University of Toronto) for providing the plasmid for Xenopus laevis calmodulin. We would like to acknowledge Prof. Lewis Kay (University of Toronto), Ranjith Muhandiram (University of Toronto), and many members from Varian Inc. and the Varian applications group (George Gray, Eriks Kupce, Mikhail Reibarkh, Bao Nguyen, and Christine Hofstetter) for their continued help. Julianne Kitevski-LeBlanc wishes to acknowledge the Natural Sciences and Engineering Research Council of Canada (NSERC) for a doctoral fellowship and RSP acknowledges NSERC, and the Ontario government for financial support through the NSERC discovery and Provincial Research Excellence Award (PREA) programs.

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Authors and Affiliations

  1. Department of Chemistry, University of Toronto, UTM, 3359 Mississauga Rd, North Mississauga, ON, L5L 1C6, Canada

    Julianne L. Kitevski-LeBlanc, Ferenc Evanics & R. Scott Prosser

  2. Department of Biochemistry, University of Toronto, Toronto, ON, M5S 1A8, Canada

    R. Scott Prosser

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  1. Julianne L. Kitevski-LeBlanc
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  2. Ferenc Evanics
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  3. R. Scott Prosser
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Correspondence to R. Scott Prosser.

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Kitevski-LeBlanc, J.L., Evanics, F. & Prosser, R.S. Approaches for the measurement of solvent exposure in proteins by 19F NMR. J Biomol NMR 45, 255–264 (2009). https://doi.org/10.1007/s10858-009-9359-2

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