What of course also could have happened, is that the Bfactor restraints are too tight, forcing, too low Bfactors for the flexible side chain. Especially after rotatable bonds, Bfactors can increase much more sharply than many of the older parameter files would allow. In this case, reducing these restraints would solve the problem.
Herman

From: CCP4 bulletin board [mailto:[log in to unmask]] On Behalf Of [log in to unmask]
Sent: Tuesday, November 20, 2012 9:14 AM
To: [log in to unmask]
Subject: Re: [ccp4bb] occupancy vs. Bfactors

Hi Robby and Grant,
Here I must second Pavel. As Grant just discovered, modern refinement programs have improved so much, that they will no longer allow to fudge a low occupancy (sharper but less density) with a high Bfactor (broader and more density), especially at higher resolutions. In this case, I would scroll down the contour level and look for alternative conformations. Even if you don't find any, the most correct approach in my view would still be to lower the occupancy, although I am personnally too lazy to do this.
 
My two cents,
Herman
 

From: CCP4 bulletin board [mailto:[log in to unmask]] On Behalf Of Robbie Joosten
Sent: Tuesday, November 20, 2012 6:46 AM
To: [log in to unmask]
Subject: Re: [ccp4bb] occupancy vs. Bfactors

Hi Grant,

This is part of the recurring side chain discussion. There is no consensus in the community about what the optimal approach is.
In your current approach you are adding a model parameter (occupancy) to improve the fit with the experimental data (remove negative difference density). You should ask yourself whether you really need to add that parameter. Are you not overfitting? Is there any clear evidence that the atoms are not always there?
The alternative model you propose (full occupancy, high B) has fewer parameters and explains more of the strucure (you account for all atoms the protein has, prior knowledge). This model probably also better reflects the uncertainty of the coordinates of the side chains involved. If your B-factor restraints are not too tight, the difference densitty should also disappear (equal explanation of the experimental data). To me that would be a better model.

HTH,
Robbie

Date: Mon, 19 Nov 2012 23:36:56 +0000
From: [log in to unmask]
Subject: [ccp4bb] occupancy vs. Bfactors
To: [log in to unmask]

Hello all,

I'm currently working on a structure which if I stub a certain side chain phenix/coot shows me a large green blob which looks strikingly similar to the side chain, when I put it in and run another refinement the blob turns red.

Basically I was just playing around and I changed the occupancy of the side chain and now there are no complaints. But I was thinking, should I haven changed the Bfactors instead? Should I have left well enough alone? If I lower the occupancy manually and do not include alternate confirmations have I introduced modelling bias?

Could someone recommend some good articles I could read on exactly how to correctly fix this problem.

Thanks,
GM