Dynamic Nuclear Polarization Study of Inhibitor Binding to the M2[subscript 18–60] Proton Transporter from Influenza A
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Griffin_Dynamic nuclear polarization.pdf
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Author(s)
Andreas, Loren
Barnes, Alexander
Chou, James J.
Caporini, Marc A.
Rosay, Melanie
Corzilius, Bjorn
Miller, Eric Alexander
Griffin, Robert Guy
Date Issued
March 2013
Journal
Biochemistry
Publisher
American Chemical Society (ACS)
Citation
Andreas, Loren B., Alexander B. Barnes, Björn Corzilius, James J. Chou, Eric A. Miller, Marc Caporini, Melanie Rosay, and Robert G. Griffin. “Dynamic Nuclear Polarization Study of Inhibitor Binding to the M2 [subscript 18–60] Proton Transporter from Influenza A .” Biochemistry 52, no. 16 (April 23, 2013): 2774–2782.
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Author's final manuscript
Abstract
We demonstrate the use of dynamic nuclear polarization (DNP) to elucidate ligand binding to a membrane protein using dipolar recoupling magic angle spinning (MAS) NMR. In particular, we detect drug binding in the proton transporter M2[subscript 18–60] from influenza A using recoupling experiments at room temperature and with cryogenic DNP. The results indicate that the pore binding site of rimantadine is correlated with previously reported widespread chemical shift changes, suggesting functional binding in the pore. Futhermore, the [superscript 15]N-labeled ammonium of rimantadine was observed near A30 [superscript 13]Cβ and G34 [superscript 13]Cα, suggesting a possible hydrogen bond to A30 carbonyl. Cryogenic DNP was required to observe the weaker external binding site(s) in a ZF-TEDOR spectrum. This approach is generally applicable, particularly for weakly bound ligands, in which case the application of MAS NMR dipolar recoupling requires the low temperatures to quench dynamic exchange processes. For the fully protonated samples investigated, we observed DNP signal enhancements of ∼10 at 400 MHz using only 4–6 mM of the polarizing agent TOTAPOL. At 600 MHz and with DNP, we measured a distance between the drug and the protein to a precision of 0.2 Å.
MIT Department
Massachusetts Institute of Technology. Department of Chemical Engineering
Massachusetts Institute of Technology. Department of Chemistry
Francis Bitter Magnet Laboratory (Massachusetts Institute of Technology)
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DOI of Published Version
https://doi.org/10.1021/bi400150x
