Current applications of 19F NMR to studies of protein structure and dynamics

Kitevski-LeBlanc, Julianne L.; Prosser, R. Scott

doi:10.1016/j.pnmrs.2011.06.003

Cited by 249 publications

(308 citation statements)

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“…In addition, 19 F is absent from virtually all naturally occurring biomolecules, thus studies of fluorinated proteins do not suffer from any background signals. Several methods to prepare 19 F-modified proteins have been described, [22][23][24][25][26][27] and recently a simple and inexpensive approach, utilizing the fluoroindole precursor for expressing 19 F-Tryptophan-containing proteins has been reported. [28] In the present work we explore 19 F PREs for extracting quantitative distance restraints, using the cyanobacterial lectin CV-N as a model system.…”

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confidence: 99%

¹⁹F Paramagnetic Relaxation Enhancement: A Valuable Tool for Distance Measurements in Proteins

Matei

Gronenborn

2015

Angewandte Chemie

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Fluorine NMR paramagnetic relaxation enhancement was evaluated as a versatile approach for extracting distance information in selectively F-labeled proteins. Proof of concept and initial applications are presented for the HIV-inactivating lectin Cyanovirin-N. Single F atoms were introduced at the 4-, 5-, 6-or 7 positions of Trp49 and the 4 position of Phe4, Phe54 and Phe80. The paramagnetic MTSL label was attached to Cys residues that were placed into the protein at positions 50 or 52. 19 F-T 2 NMR spectra with different relaxation delays were recorded and the transverse 19 F-PRE rate, 19 F-Γ 2 , was used to determine the average distance between the F nucleus and the paramagnetic center. Our data show that experimental 19 F PRE based distances correspond to ~0.93 of the 1 H N -PRE distances, in perfect agreement with the gyromagnetic γ 19 F/γ 1 H ratio, thereby demonstrating that 19 F PREs are excellent alternative parameters for quantitative distance measurements in selectively F-labeled proteins. Keywords fluorine; paramagnetic; NMR; distance; protein In Paramagnetic Relaxation Enhancement (PRE) NMR experiments the increase in longitudinal or transverse relaxation rates, induced by the presence of a paramagnetic center, is measured. The effect is distance dependent, and, as a result, information on the distance between the paramagnetic moiety to the measured nuclei is obtained.In biomolecular NMR, PREs are increasingly used for providing long-range distance information that complements nuclear Overhauser effect (NOE)-derived, short (≤ 5 Å) interproton distance restraints. In cases where NOE data were found limited, distances derived from PREs provided valuable supplemental restraints and permitted either characterization of the global folds for some proteins, [1,2] or delineate structural properties of disorder proteins. [3,4] Correspondence to: Angela M. Gronenborn, amg100@pitt.edu. Supporting information for this article is given via a link at the end of the document. HHS Public AccessAuthor manuscript Angew Chem Int Ed Engl. Author manuscript; available in PMC 2017 January 04. Author Manuscript Author ManuscriptAuthor Manuscript Author ManuscriptThe most widely used spin label, originally developed for EPR studies, [5,6] is the nitroxide radical MTSL (1-oxyl-2,2,5,5-tetramethyl-Δ 3 -pyrroline-3-methyl)methanethiosulfonate. [7,8] It enhances the transverse relaxation rate (R 2 ) of protons and is introduced into proteins via site specific labeling of cysteine residues. [9][10][11] PRE measurements in proteins commonly monitor 1 H line broadening or intensity attenuation of proton resonances, caused by R 2 relaxation rate enhancements, and these are used to derive distances, with an accessible distance range for the MTSL spin label to protons of 13-25 Å. Resonances of residue in close proximity to the spin label are broadened beyond detection, while for long distances, the effect becomes undetectable.

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¹⁹F Paramagnetic Relaxation Enhancement: A Valuable Tool for Distance Measurements in Proteins

Matei

Gronenborn

2015

Angewandte Chemie

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“…19 F-NMR has been used extensively in studies of protein structure and interactions [24,25], and proves to be a valuable tool in FBDD campaigns as well as in probing the binding of other ligands relevant to such efforts.…”

Section: Characterizing Ligand Binding By 19 F-nmrmentioning

confidence: 99%

Applications of 19F-NMR in Fragment-Based Drug Discovery

et al. 2016

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19 F-NMR has proved to be a valuable tool in fragment-based drug discovery. Its applications include screening libraries of fluorinated fragments, assessing competition among elaborated fragments and identifying the binding poses of promising hits. By observing fluorine in both the ligand and the target protein, useful information can be obtained on not only the binding pose but also the dynamics of ligand-protein interactions. These applications of 19 F-NMR will be illustrated in this review with studies from our fragment-based drug discovery campaigns against protein targets in parasitic and infectious diseases.

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“…Thus, no special precautions are needed to remove buffer and additive signal intensity from the spectra. The van der Waals radius of the 19 F atom (1.47 Å ) lies between those of hydrogen (1.2 Å ) and oxygen (1.52 Å ), and strategic substitution of 19 F atoms for hydrogens, hydroxyl groups, or carbonyl oxygens in biological molecules is considered weakly perturbing and often has little effect on a protein's biological activity (Campos-Olivas, Aziz, Helms, Evans, & Gronenborn, 2002;Danielson & Falke, 1996;Gerig, 1994;Kitevski-Leblanc & Prosser, 2012).…”

Section: Introductionmentioning

confidence: 99%

“…For 40 years now, the 19 F atom has been exploited in biological NMR (Arseniev et al, 1986;Chaiken, Freedman, Lyerla, & Cohen, 1973), and several reviews have covered the field over the last two decades (Danielson & Falke, 1996;Didenko, Liu, Horst, Stevens, & Wüthrich, 2013;Gerig, 1994;Kitevski-Leblanc & Prosser, 2012;Marsh & Suzuki, 2014). Indeed, the 19 F atom has been used as a molecular probe to gain insight into protein and peptide structure (Danielson & Falke, 1996;Gerig, 1994;Kawahara et al, 2012;Liu, Horst, Katritch, Stevens, & Wuthrich, 2012), protein-ligand interactions (Campos-Olivas et al, 2002;Danielson & Falke, 1996;Gerig, 1994; Kitevski-Leblanc & Prosser, 2012;Luck & Falke, 1991;Rydzik et al, 2014), protein unfolding (Arseniev et al, 1986;Chaiken et al, 1973;Khan, Kuprov, Craggs, Hore, & Jackson, 2006;Kitevski-Leblanc, Hoang, Thach, Larda, & Prosser, 2013), protein aggregation (Danielson & Falke, 1996;Didenko et al, 2013;Gerig, 1994;Kitevski-Leblanc & Prosser, 2012;Li et al, 2009;Marsh & Suzuki, 2014;Suzuki, Brender, Hartman, Ramamoorthy, & Marsh, 2012), and protein dynamics (Fischer et al, 2003), clearly demonstrating the power and versatility of the fluorine probe for NMR.…”

Section: Introductionmentioning

confidence: 99%

19F-Modified Proteins and 19F-Containing Ligands as Tools in Solution NMR Studies of Protein Interactions

Sharaf

Gronenborn

2015

Methods in Enzymology

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Current applications of 19F NMR to studies of protein structure and dynamics

Cited by 249 publications

References 145 publications

¹⁹F Paramagnetic Relaxation Enhancement: A Valuable Tool for Distance Measurements in Proteins

¹⁹F Paramagnetic Relaxation Enhancement: A Valuable Tool for Distance Measurements in Proteins

Applications of 19F-NMR in Fragment-Based Drug Discovery

19F-Modified Proteins and 19F-Containing Ligands as Tools in Solution NMR Studies of Protein Interactions

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Current applications of 19F NMR to studies of protein structure and dynamics

Cited by 249 publications

References 145 publications

19F Paramagnetic Relaxation Enhancement: A Valuable Tool for Distance Measurements in Proteins

19F Paramagnetic Relaxation Enhancement: A Valuable Tool for Distance Measurements in Proteins

Applications of 19F-NMR in Fragment-Based Drug Discovery

19F-Modified Proteins and 19F-Containing Ligands as Tools in Solution NMR Studies of Protein Interactions

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¹⁹F Paramagnetic Relaxation Enhancement: A Valuable Tool for Distance Measurements in Proteins

¹⁹F Paramagnetic Relaxation Enhancement: A Valuable Tool for Distance Measurements in Proteins