On Tue, 7 Jan 2014, Jan Zalesak wrote:

I did not find a reliable way to follow the distance calculations from
intensities/volumes of the peaks.

I have tried 3 steps (lets say I have overlap of 3Adenine H6-H2' peak with 2Ura H6-H2'
peak)
1) I have put 3Adenine H6-H2' peak
2) I have manually added the second (2Ura H6-H2' peak) - "crosses" - on one peak
(manually).
3) I have deleted (2Ura H6-H2' peak) and filled the assignment of it to the assignment
of the first peak (so i have ambiguous restraint)

After every step I check the peak intensities, peak volumes) and I calculate the
distances.
But surprisingly the values (all of them - intensities, peak volumes, calculated
distances) do not change for any of the two resonances.

If your peak created in step 2 is close to the first peak then it will pick up the same intensity value & volume because it's looking at the same bit of the spectrum.

When you create a distance restraint from a peak in analysis, you simply map the intensity or volume to a distance by the algorithm you choose (1/r6, bins, etc.), so that won't change from situation to situation you describe. What that then means to the structure calculation program depends on the structure calculation program - for ambiguous restraints you want to use one that treats the distance to intensity mapping as "sum" in ARIA/CNS terminology.

You should use the third case you have (ambiguous) - you might want to reduce the weighting of the contribution of the NOE spectrum in the chemical shift list if you're going to manually assign multiple possibilities to an ambiguous peak that might be a little off the exact shift of each possible contributor. You could leave ambiguous peaks (partially) unassigned and use "make shift match restraints" to avoid this issue.

Is there a way to handle this?

Or is there a way to do it properly, but in a way program will understand that there are
two peaks and the distance I want him to calculate should be smaller than if there was a
single peak?

I think you misunderstood the intensity->distance mapping process.

Can I for example change the peak boxes manually (or at least visualize them?)

It would be nice to have peak "footprints" that would allow the use of overlapped NOE peaks more completely in the creation of ambiguous distance restraints - currently, really overlapped peaks just have to be discarded.

--
Dr. Brian O. Smith --------------------------- Brian Smith at glasgow ac uk
Institute of Molecular, Cell and Systems Biology & School of Life Sciences,
          College of Medical, Veterinary & Life Sciences,
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