Dear Kendall,
I would suggest you to run a simulated annealing omit map around the
region you are interested.
The model bias will be reduced and you will get a more clear answer.
Ciao,
Joao
Joao M. Dias
Ollmann Saphire Lab
The Scripps Research Institute
10550 North Torrey Pines Rd. IMM-2
La Jolla, CA 92037 USA
tel (858)784-8925
On May 24, 2007, at 9:57 AM, Kendall Nettles wrote:
I’d like help in interpreting some mystery density in a structure.
I’m writing a paper about soaking the apo-estrogen receptor with
different ligands. The apo structure is already released, as pdb
code 2B23. The question is whether there is a mystery molecule in
the pocket of the apo receptor. If you superimpose 3ERD, you can
see where the ligand binds. The problem is that with some maps the
pocket appears completely empty, and with others, there appears to
be something there. Protein looks essentially identical with the
different maps. We have used a few different approaches to identify
the compound with LC-MS, and are pretty sure there is nothing
there. For example, we can bind our protein to beads, soak it with
estradiol, wash extensively, elute with organic solvent and find a
great peak for estradiol, but nothing for the apo protein. We have
also tried non-denaturing MS.
If you look at the 2mFo-DFc map from EDS in Coot or CCP4mg, you see
mystery density in the pocket. If I use the MTZ, you see density
in Coot, but not CCP4MG. I then downloaded the structure factors
from the PDB and made an MTZ. The map in CCP4MG shows some density,
but much less that with the map from EDS. When I used FFT to make a
2mF1-1nF2 map, there is no mystery density in either CCP4MG or coot.
I was ready to submit the manuscript with a picture of the mystery
density, but now I’m not sure if that is appropriate. Any
suggestions, as far as how to interpret this mystery density would
be greatly appreciated.
Best Regards,
Kendall
--
Kendall W. Nettles, PhD
Assistant Professor
Department of Cancer Biology
The Scripps Research Institute
5353 Parkside Dr.
Jupiter Fl 33458
office 561-799-8851
fax 561-799-8805
cell 561-306-7566
