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Off-resonance rotating-frame amide proton spin relaxation experiments measuring microsecond chemical exchange in proteins

Lundström, Patrik LU and Akke, Mikael LU (2005) In Journal of Biomolecular NMR 32(2). p.163-173
Abstract
NMR spin relaxation in the rotating frame (R-1 rho) is a unique method for atomic-resolution characterization of conformational ( chemical) exchange processes occurring on the microsecond time scale. Here, we use amide H-1 off-resonance R-1 rho relaxation experiments to determine exchange parameters for processes that are significantly faster than those that can be probed using N-15 or C-13 relaxation. The new pulse sequence is validated using the E140Q mutant of the C-terminal domain of calmodulin, which exhibits significant conformational exchange contributions to the transverse relaxation rates. The H-1 off-resonance R-1 rho data sample the entire relaxation dispersion profiles for the large majority of residues in this protein, which... (More)
NMR spin relaxation in the rotating frame (R-1 rho) is a unique method for atomic-resolution characterization of conformational ( chemical) exchange processes occurring on the microsecond time scale. Here, we use amide H-1 off-resonance R-1 rho relaxation experiments to determine exchange parameters for processes that are significantly faster than those that can be probed using N-15 or C-13 relaxation. The new pulse sequence is validated using the E140Q mutant of the C-terminal domain of calmodulin, which exhibits significant conformational exchange contributions to the transverse relaxation rates. The H-1 off-resonance R-1 rho data sample the entire relaxation dispersion profiles for the large majority of residues in this protein, which exchanges between conformations with a time constant of approximately 20 mu s. This is in contrast to the case for N-15, where additional laboratory-frame relaxation data are required to determine the exchange parameters reliably. Experiments were performed on uniformly N-15-enriched samples that were either highly enriched in H-2 or fully protonated. In the latter case, dipolar cross-relaxation with aliphatic protons were effectively decoupled to first order using a selective inversion pulse. Deuterated and protonated samples gave the same results, within experimental errors. The use of deuterated samples increases the sensitivity towards exchange contributions to the H-1 transverse relaxation rates, since dipolar relaxation is greatly reduced. The exchange correlation times determined from the present H-1 off-resonance R-1 rho experiments are in excellent agreement with those determined previously using a combination of N-15 laboratory-frame and off-resonance R-1 rho relaxation data, with average values of <tau(ex)> = 19 +/- 7 and 21 +/- 3 mu s, respectively. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Journal of Biomolecular NMR
volume
32
issue
2
pages
163 - 173
publisher
Springer
external identifiers
  • pmid:16034667
  • wos:000230694100006
  • scopus:23344439330
ISSN
1573-5001
DOI
10.1007/s10858-005-5027-3
language
English
LU publication?
yes
id
69c2f928-2f2a-4bd1-af31-30cdbf7e4b8e (old id 151504)
date added to LUP
2007-06-29 13:15:16
date last changed
2017-08-06 03:37:53
@article{69c2f928-2f2a-4bd1-af31-30cdbf7e4b8e,
  abstract     = {NMR spin relaxation in the rotating frame (R-1 rho) is a unique method for atomic-resolution characterization of conformational ( chemical) exchange processes occurring on the microsecond time scale. Here, we use amide H-1 off-resonance R-1 rho relaxation experiments to determine exchange parameters for processes that are significantly faster than those that can be probed using N-15 or C-13 relaxation. The new pulse sequence is validated using the E140Q mutant of the C-terminal domain of calmodulin, which exhibits significant conformational exchange contributions to the transverse relaxation rates. The H-1 off-resonance R-1 rho data sample the entire relaxation dispersion profiles for the large majority of residues in this protein, which exchanges between conformations with a time constant of approximately 20 mu s. This is in contrast to the case for N-15, where additional laboratory-frame relaxation data are required to determine the exchange parameters reliably. Experiments were performed on uniformly N-15-enriched samples that were either highly enriched in H-2 or fully protonated. In the latter case, dipolar cross-relaxation with aliphatic protons were effectively decoupled to first order using a selective inversion pulse. Deuterated and protonated samples gave the same results, within experimental errors. The use of deuterated samples increases the sensitivity towards exchange contributions to the H-1 transverse relaxation rates, since dipolar relaxation is greatly reduced. The exchange correlation times determined from the present H-1 off-resonance R-1 rho experiments are in excellent agreement with those determined previously using a combination of N-15 laboratory-frame and off-resonance R-1 rho relaxation data, with average values of &lt;tau(ex)&gt; = 19 +/- 7 and 21 +/- 3 mu s, respectively.},
  author       = {Lundström, Patrik and Akke, Mikael},
  issn         = {1573-5001},
  language     = {eng},
  number       = {2},
  pages        = {163--173},
  publisher    = {Springer},
  series       = {Journal of Biomolecular NMR},
  title        = {Off-resonance rotating-frame amide proton spin relaxation experiments measuring microsecond chemical exchange in proteins},
  url          = {http://dx.doi.org/10.1007/s10858-005-5027-3},
  volume       = {32},
  year         = {2005},
}