UCLA-DOE Biosciences

For 15N CPMG relaxation dispersion, data should be collected using the CW CPMG experiment from
Hansen, Vallurupalli, and Kay, J. Phys. Chem. B 112, 5898-5904 (2008)
Our version of the pulse program is called 15N_R2cpmg_HC_<xwin/TS3>.rp

The CPMG experiments should be processed with nmrPipe. An initial processing script can be generated from within Topspin3 using the au program convert2pipe.ecj33. You can use standard processing initially.

Note: all of the files and scripts discussed below will be placed in and run in the Main NMR Directory. This is the Topspin directory that contains all the data for the series you’re analyzing (ie the <expt name> directory).

nmrPipe has tools to analyze 2D relaxation data. You can find a description of it on the nmrPipe website. Follow the instructions for using the GM window function during nmrPipe processing. Then use proc.com, fid.com, and ft2.com to process all spectra in the series. tauList  and dirList inside proc.com have to be modified to include your specific CPMG “frequencies” and the directories that correspond to those frequencies. Run by typing ./proc.com. After that, pick and assign peaks in one of the spectra (using nmrPipe). nmrPipe calls the saved peak list “test.tab” by default. Rename this file relax.master.tab, and place it in the Main NMR Directory. Then run fit.com (by typing ./fit.com). It can take awhile to run. Just be patient and wait until the little blue window disappears and you get the prompt back. It will generate two new files called axt.tab and nlin.tab.

I have written Python scripts to take all the data from nlin.tab (or axt.tab), and order it appropriately for further analysis. Place the script order_CPMG_intensities_nmrpipe.py in the Main NMR Directory. Inside the script, modify the “offsets” list, which contains the CPMG frequencies – it should have the same values as tauList inside proc.com. Create a directory called output_intensities, then type ./order_CPMG_intensities_nmrpipe.py to run it. This will create a separate file for each residue (in the directory output_intensities). These files have 3 columns: 1) The CPMG frequency, 2) The intensity, 3) An error estimate. Next place the script order_CPMG_R2_nmrpipe.py in the Main NMR Directory, and modify the “offsets” list as in order_CPMG_intensities_nmrpipe.py. Create a directory called output_R2, then type ./order_CPMG_R2_nmrpipe.py to run it. This also creates separate files for each residue in the directory output_R2. These files also have 3 columns: 1) The CPMG frequency, 2) The R2 value, 3) An error estimate (the error estimate is not done correctly).

Run the scripts graph_intensities.py and graph_R2.py to see graphs of the peak intensities/R2 vs. CPMG frequency for each residue. These scripts generate the graph for one residue at a time. When you click the graph for the 1st residue away, the next one will appear, and so one. This is just a rough way to visualize the data. If some residues look promising, you can proceed with analysis using ChemEx or other software.


Example scripts are in the directory /home/peterson/NMR/CPMG_analysis

fid_v33.com      (initial nmrPipe processing script)
proc.com            (proc.com is a script that contains the CPMG “frequencies” and a list)
fid.com               (of the directories containing the experiments done using those frequencies)
ft2.com               (fid.com and ft2.com are nmrPipe processing scripts that use proc.com as input)
fit.com                (script that fits intensities from all spectra in the group to a decaying exponential)
order_CPMG_intensities_nmrpipe.py   (Python script that orders the intensities for each residue in a separate file)
order_CPMG_R2_nmrpipe.py                 (Python script that orders the R2 values for each residue in a separate file)
graph_intensities.py     (Python script that graphs the intensity vs. CPMG frequency for each residue separately)
graph_R2.py                   (Python script that graphs the R2 value vs. CPMG frequency for each residue separately)


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