10.1021/acs.jpclett.7b02709.s001
Dwaipayan Mukhopadhyay
Dwaipayan
Mukhopadhyay
Philippe S. Nadaud
Philippe S.
Nadaud
Matthew D. Shannon
Matthew D.
Shannon
Christopher P. Jaroniec
Christopher P.
Jaroniec
Rapid Quantitative Measurements of Paramagnetic Relaxation
Enhancements in Cu(II)-Tagged Proteins by Proton-Detected Solid-State
NMR Spectroscopy
American Chemical Society
2017
PRE
proton-detected 2 D
3 D correlation spectra
13 C
28C
GB
2 H
experiment time
Proton-Detected Solid-State NMR Spectroscopy
natively diamagnetic proteins
binding tags
Paramagnetic Relaxation Enhancements
H 2 O
electron relaxation time
1 H
Cu
backbone amide 15 N
Rapid Quantitative Measurements
relaxation enhancements
structure determination
15 N-labeled protein back-exchanged
2017-11-17 00:00:00
Journal contribution
https://acs.figshare.com/articles/journal_contribution/Rapid_Quantitative_Measurements_of_Paramagnetic_Relaxation_Enhancements_in_Cu_II_-Tagged_Proteins_by_Proton-Detected_Solid-State_NMR_Spectroscopy/5616562
We
demonstrate rapid quantitative measurements of site-resolved
paramagnetic relaxation enhancements (PREs), which are a source of
valuable structural restraints corresponding to electron–nucleus
distances in the ∼10–20 Å regime, in solid-state
nuclear magnetic resonance (NMR) spectra of proteins containing covalent
Cu<sup>2+</sup>-binding tags. Specifically, using protein GB1 K28C-EDTA-Cu<sup>2+</sup> mutant as a model, we show the determination of backbone
amide <sup>15</sup>N longitudinal and <sup>1</sup>H transverse PREs
within a few hours of experiment time based on proton-detected 2D
or 3D correlation spectra recorded with magic-angle spinning frequencies
≥ ∼ 60 kHz for samples containing ∼10–50
nanomoles of <sup>2</sup>H,<sup>13</sup>C,<sup>15</sup>N-labeled protein
back-exchanged in H<sub>2</sub>O. Additionally, we show that the electron
relaxation time for the Cu<sup>2+</sup> center, needed to convert
PREs into distances, can be estimated directly from the experimental
data. Altogether, these results are important for establishing solid-state
NMR based on paramagnetic-tagging as a routine tool for structure
determination of natively diamagnetic proteins.