Dear colleagues,
First of all, I would like to thank to all who responded! I have
selected some answers and my reactions to sum up:
1) Kay Diedrichs - try a Polder map: clear density for the ligand at 3
sigma appeared.
2) JR Helliwell - build first five atoms and check the density now (read
Minichino et al Acta Cryst D 2002): clear difference density for another
eight atoms with correct geometry appeared. I did not do this
previously, because these five atoms are half of a compact ring. I was
sure that I must see the WHOLE ring in case of the right observation.
These two checks together with fine refinement of the ligand with
reduced occupancy convinced me to use the dataset. Initially, I thought
the dataset is useless. Take-home message for me:
- Do not give up too early and try more calculation procedures with
every dataset you have.
- Go through you old non-complex data and try new procedures.
There are some other calculations to perform (e.g. Fo,soak - Fo,native
map calculation). And although I am not a big fan of huge data storage,
I will deposit the diffraction images together with the coordinates
especially for this structure!
Once again, thanks for your help!
Petr
Dne 08.02.2017 v 9:05 Petr Kolenko napsal(a):
Dear colleagues,
we have a dataset with potential enzyme:ligand complex at 2.2 AA
resolution. The ligand is very good substrate for the enzyme, we used
soaking. We do not see the ligand in the regular difference electron
density, only five out of twenty atoms. However, the ligand placed at
the active site (model used from structure of a mutant variant) is
refined well, giving no negative peaks in difference electron density
map and nice observed electron density. I have calculated composit
omit map with annealing in Phenix (input model did not contain the
ligand) and the electron density for the ligand is there.
I have my own opinion, but we are desperate to obtain such data (more
than 40 crystals already tested). My question is, would this be proof
of presence of the ligand with reduced occupancy? Will this map
convince the reviewers? Is there any other way to validate presence of
the ligand?
Best regards,
Petr
