Thanks for the replies. Yuri, yes, those were my exact concerns. This
is a molecular replacement structure, but that was done with an apo
structure, so I am certain the metal is there. My resolution is around
2.3 angstroms, which I think is OK for determining geometry.
I have done omit maps as well as simply several rounds of refinement
with no metal present. It always comes back, and has a certain shape
(tetrahedral).
I just was concerned that I was forcing a geometry by not specifying
nickel properly in my pdb file. I didn't know right off how one might
specify different oxidation states in a pdb file (say Nickel I vs Nickel
II; that is not what I am trying to do, but it is something I'd be
curious about). I haven't worked with a lot of metalloenzymes, so I
just wanted to be sure. I think you are right, and I am doing things
properly (the programs are of course very good).
Thanks again
Dave
On 5/12/2012 2:49 PM, Yuri Pompeu wrote:
Hi Dave,
I sounds to me like you are worried about 2 separate things here.
A: Am I affecting the geometry of the coordination sites with a restraint file
that is innacurate?
B:Are my electron density maps biased, and what I am seeing is not really there?
AFAIU, if you have a restraint file that is innacurate, lets say it is defining the
metal/ ligand angles to be those of a tetrahedron, that would influence the position of
your atoms after refinement as the program will try to "obey" the restraint
file.
The electron density maps, however do not directly take into account your
restraints file.
With that being said, model bias can be a problem, yes. This is dependent on
many factors, and if you have obtained your phases through a molecular
replacement solution rather than experimentally (MAD, SAD, etc..) your maps
will be particularly susceptible to bias. And if your dealing with a low
resolution data set this can become even more of a problem (you dont mention
your resolution). If you are working with a 1.2A data set, I would not lose
sleep over it. People have spent many hours of thinking and programming to
develop ways of eliminating model bias and many programs can calculate electron
density maps in a way that your bias is minimized. Always check your difference
map (mFo-Dfc), you can calculate omit maps, averaged kick maps, and in the case
of metals even an anomalous maps sometimes. All of these would help you put
your mind at ease.
Hope this helps
