Dear Dominik,
I do not see any problems with a mosaicity of 0.48°. Also the Wilson B-factor of 30.34 of your data is ok, indicating no problems with the processing. Your problem is that the average REFINED b-factor is much higher than your "observed" average b. The first thing I would do is to calculate the average b-factor for your protein only. Adding a lot of high b-factor water molecules can increase your average b-factor significantly. If the average- and wilson B are then in the same ballpark, I would not worry too much and rahter have a look at my waters to see if some of them could be removed. If the descrepancy is still there for the protein only, I would look at the way the b-factors are refined (individual, grouped, TLS, weights, restraints etc.) to see if there is a problem somewhere.
Best regards,
Herman
-----Original Message-----
From: CCP4 bulletin board [mailto:[log in to unmask]] On Behalf Of Dominik Possner
Sent: Thursday, October 08, 2009 5:59 PM
To: [log in to unmask]
Subject: [ccp4bb] temperature factors, mosaicity and wilson plot
Hi,
I recently solved the structure of a mutant protein that contains a single amino acid substitution.
Here are some data about the mutant:
resolution: 2.65 A
completeness: 99.9 % (100.0 %)
Rwork/free: 22.3/27.8 %
Rsym 15.6 (47.7) %
avg. mosaicity: 0.48°
now whats confusing me:
Wilson b-factor: 30.34 A^2
Overall mean b-factor: 55.92 A^2
My questions are the following:
Where could the large difference of the b-factors come from? As noted, the crystal shows a mosaicity of avg. 0.48°.
Is there a way I could evaluate whether the high b-factors are completely explained by the high mosaicity? What other factors could explain the high b-factors? Does the b-factor correlate that clearly with the resolution?
I used XDS for data processing and REFMAC for refinement. Does either of these programs consider the mosaicity so that i could exclude that the b-factors origin from the mosaicity?
Thanks,
Dominik
|