3D examples in Bruker format (courtesy of the University of Florence)

Prof. Mario Piccioli (CERM, University of Florence) has generously provided an assorted set of 3D spectra of a metalloprotein (the reduced form of copper, zinc superoxide dismutase from Salmonella enterica). They are listed at the bottom of this page. The NMR resonances have been almost completely assigned and deposited at the BMRB archive. Other authors are: Beatriz Jimenez, Mirko Mori, Andrea Battistoni and Marco Sette. I want to thank them all for the very nice spectra. Please note that they are not responsible for this web site and for the way the files have been packaged.

You have the opportunity to learn and process these 3D data sets with iNMR. The data files have been stripped down, but the “ser” files are intact, so you can open them with other programs like NMRPipe. If you open the spectra with iNMR, it will automatically process them. The given processing parameters are merely indicative. They don't come from the cited owners and authors. This is work in progress and better parameters can be enclosed in future.. If you are curious about which processing iNMR performs on each spectrum, select the command “Edit/Copy.../Processing”, then paste into an empty page (or into the spectrum). The list of performed actions will appear. You are free to reload the FID (command: “File/Reload”) and apply the processing of your choice.

3D processing is RAM-thirsty. If your computer is forced to “swap” memory pages, then everything will become painfully slow. It can be avoided by quitting the unused applications. A more essential rule is to never combine zero-filling (or LP filling) with the hyper-complex option. Beware that iNMR by default turns on both zero-filling and the hyper-complex option! You must disable one of them manually (unless you keep the processing proposed hereby, of course). If, instead, your computer has 4 GB of RAM and is fast enough, you can use both zero-filling and the hyper-complex option.

The purpose of this site is also to demonstrate the 3D capabilities of iNMR and iNMR reader. Both programs can be freely downloaded. The processing parameters I am proposing for these five experiments can be used for different spectra acquired with the same pulse sequences. Weighting functions and other FT options are the default choices provided by iNMR 3.3.8. Scales are referenced according to the “O1” parameter found into the Bruker files. Other treatments are: digital water suppression and NON-polynomial base-line corrections (performed along the direct dimension only). In all the examples provided, iNMR over-estimates the baseline to subtract and therefore leaves negative traces. Sometimes the negative peaks are hidden (the negative contour levels are not shown), but you may choose to show them. Alternatively, you can perform a further processing with the console command "noneg()" which cancels all the negative elements of a matrix.

To select the planes for observation, go to the last panel of the Scale dialog (Cmd-G). The f1-f3 are the most informative planes. To swap the x and y axes there is a shortcut that bypasses the dialog: press T (uppercase) on the keyboard. The keys r and t (lowercase) let you move across the slices of the 3D cube.

You will find the phase already corrected but it's not clear, from the examples, how the corrections were determined. Here's the recipe.
Extract the first plane (create a single horizontal mark as low as possible, then press Cmd-E). Repeat the same operation to extract the first row. Fourier Transform. Correct the phase (a zero order correction is enough). Close the extract twice (or Reload the whole data set). Perform the three FTs (with the hypercomplex option along the indirect dimensions) and with minimal zero-filling. Remember that the number of points along the indirect dimension is halved because they are phase-sensitive spectra. For example: if you see 48 increments along the 15-N dimension, they will become 24 points before FT, therefore a size of 32 does not truncate your data: it's a minimal and effective zero-filling. Once you have arrived into the frequency domain, select the side containing the f1 and f3 dimensions. Browse through the planes until you find the plane with the maximum number of peaks. Now correct the phase along both directions, if needed. First Order corrections may be necessary. When done, transpose the cube to show the f2-f3 planes and check the phase. Correct it, if required. When you have arrived at the best correction parameters, reload the FID and reprocess it. This time uncheck the hyper-phase option and zero-fill as much as you like.

Different pulse programs and different instruments will require different processing parameters. We'll try, if possible, to add other sets of examples. You can contribute.

Read also: Rational 3-D Processing Step-By-Step.


 
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