Paramagnetic Constraints in Structure Determination: Difference between revisions
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== '''Introduction''' == | == '''Introduction''' == | ||
The collection of paramagnetic constraint data and the use of those data as a part of a structure determination is fairly straight forward. RDCs can be collected using either the J-modulation or HSQC-TROSY methods described in the RDC section. Paramagnetic relaxation enhancements (PREs) are collected based on attenuation of signal intensity in HSQC or TROSY spectra. When Curie contributions from paramagnetic metals dominate it will be convenient to make measurements at multiple field strengths as the effects are field squared dependent and field variation provides a useful way to probe different distance ranges. We have access to spectrometers operating from 600 to 900 MHz. Pseudocontact shifts (PCSs) are measured by comparison of HSQC (or TROSY) cross-peak positions in diamagnetic (La<sup>3+</sup>) and paramagnetic complexes (Dy<sup>3+</sup> or Tb<sup>3+</sup>). Pairing of shifted and non-shifted peaks is facilitated by the fact that shifts in in both <sup>1</sup>H and <sup>15</sup>N dimensions are nearly equal on the ppm scale and are therefore connected by diagonal lines. Paramagnetic relaxation interferences (PRIs) produce differential effects on the α and β cross peaks of coupled HSQC spectra and the cross-correlation effects can be measured using experiments that we have developed for the measurement of correlation times from CSA/DD interference (Ref. 1). Integration of these data into structure characterization protocols in the NESG is accomplished using programs such as XPLOR-NIH (Ref. 2) or REDCAT (Ref. | The collection of paramagnetic constraint data and the use of those data as a part of a structure determination is fairly straight forward. RDCs can be collected using either the J-modulation or HSQC-TROSY methods described in the RDC section. Paramagnetic relaxation enhancements (PREs) are collected based on attenuation of signal intensity in HSQC or TROSY spectra. When Curie contributions from paramagnetic metals dominate it will be convenient to make measurements at multiple field strengths as the effects are field squared dependent and field variation provides a useful way to probe different distance ranges. We have access to spectrometers operating from 600 to 900 MHz. Pseudocontact shifts (PCSs) are measured by comparison of HSQC (or TROSY) cross-peak positions in diamagnetic (La<sup>3+</sup>) and paramagnetic complexes (Dy<sup>3+</sup> or Tb<sup>3+</sup>). Pairing of shifted and non-shifted peaks is facilitated by the fact that shifts in in both <sup>1</sup>H and <sup>15</sup>N dimensions are nearly equal on the ppm scale and are therefore connected by diagonal lines. Paramagnetic relaxation interferences (PRIs) produce differential effects on the α and β cross peaks of coupled HSQC spectra and the cross-correlation effects can be measured using experiments that we have developed for the measurement of correlation times from CSA/DD interference (Ref. 1). Integration of these data into structure characterization protocols in the NESG is accomplished using programs such as XPLOR-NIH (Ref. 2,3) or REDCAT (Ref. 4). [[CYANA|CYANA]] can also accommodate pseudocontact shifts. | ||
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== '''Protocols''' == | == '''Protocols''' == | ||
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1. Liu, Y.Z., and Prestegard, J.H. (2008) Direct measurement of dipole-dipole/CSA cross-correlated relaxation by a constant-time experiment. ''J. Magn. Reson. 193'', 23-31. | 1. Liu, Y.Z., and Prestegard, J.H. (2008) Direct measurement of dipole-dipole/CSA cross-correlated relaxation by a constant-time experiment. ''J. Magn. Reson. 193'', 23-31. | ||
2. Schwieters, C.D., Kuszewski, J.J., and Clore, G.M. (2006). Using Xplor-NIH for NMR molecular structure determination. ''Prog. NMR Spect. 48'', 47-62. | 2. Schwieters, C.D., Kuszewski, J.J., and Clore, G.M. (2006). Using Xplor-NIH for NMR molecular structure determination. ''Prog. NMR Spect. 48'', 47-62.<br> | ||
3. Banci, L., Bertini, I., Cavallaro, G., Giachetti, A., Luchinat, C., and Parigi, G. (2004) Paramagnetism-based restraints for Xplor-NIH. ''J. Biomol. NMR 28'', 249-261. | |||
4. Valafar, H., and Prestegard, J.H. (2004) REDCAT: a residual dipolar coupling analysis tool. ''J. Magn. Reson. 167'', 228-241 | |||
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-- Prestegard Lab - Nov 2009 | -- Prestegard Lab - Nov 2009 | ||
Revision as of 20:09, 13 November 2009
Introduction
The collection of paramagnetic constraint data and the use of those data as a part of a structure determination is fairly straight forward. RDCs can be collected using either the J-modulation or HSQC-TROSY methods described in the RDC section. Paramagnetic relaxation enhancements (PREs) are collected based on attenuation of signal intensity in HSQC or TROSY spectra. When Curie contributions from paramagnetic metals dominate it will be convenient to make measurements at multiple field strengths as the effects are field squared dependent and field variation provides a useful way to probe different distance ranges. We have access to spectrometers operating from 600 to 900 MHz. Pseudocontact shifts (PCSs) are measured by comparison of HSQC (or TROSY) cross-peak positions in diamagnetic (La3+) and paramagnetic complexes (Dy3+ or Tb3+). Pairing of shifted and non-shifted peaks is facilitated by the fact that shifts in in both 1H and 15N dimensions are nearly equal on the ppm scale and are therefore connected by diagonal lines. Paramagnetic relaxation interferences (PRIs) produce differential effects on the α and β cross peaks of coupled HSQC spectra and the cross-correlation effects can be measured using experiments that we have developed for the measurement of correlation times from CSA/DD interference (Ref. 1). Integration of these data into structure characterization protocols in the NESG is accomplished using programs such as XPLOR-NIH (Ref. 2,3) or REDCAT (Ref. 4). CYANA can also accommodate pseudocontact shifts.
Protocols
References
1. Liu, Y.Z., and Prestegard, J.H. (2008) Direct measurement of dipole-dipole/CSA cross-correlated relaxation by a constant-time experiment. J. Magn. Reson. 193, 23-31.
2. Schwieters, C.D., Kuszewski, J.J., and Clore, G.M. (2006). Using Xplor-NIH for NMR molecular structure determination. Prog. NMR Spect. 48, 47-62.
3. Banci, L., Bertini, I., Cavallaro, G., Giachetti, A., Luchinat, C., and Parigi, G. (2004) Paramagnetism-based restraints for Xplor-NIH. J. Biomol. NMR 28, 249-261.
4. Valafar, H., and Prestegard, J.H. (2004) REDCAT: a residual dipolar coupling analysis tool. J. Magn. Reson. 167, 228-241
-- Prestegard Lab - Nov 2009