Dear Jiamu Du,

what were the exact concentrations (Molar values please) of protein and 
peptide in the co-crystallization experiment? This may help in 
estimating the (possible) occupancy of your peptide.

Jeroen.



JDwrote:
> Does anyone know a program can perform the ocupancy refinement?
> Or we always only refine B factor to reflect the occupancy?
>  
> Thanks
>
>  
> On 4/30/07, *Eleanor Dodson* <[log in to unmask] 
> <mailto:[log in to unmask]>> wrote:
>
>     Well - it is extremely likely that the peptide is partially
>     occupied and
>     the occupancy may well be < 0.5..
>
>     But at this resolution you are going to have great difficulty deciding
>     whether you should have
>     Occ=1.0 <B = 130>
>
>     Occ = 0.5  <B = 100>
>
>     Occ = 0.33  <B = ??? 80???>
>
>     As your Rfactors show it makes very little difference to any scoring
>     system..
>
>     You can look at difference maps and try to see if one looks
>     flatter than
>     the other ..
>
>     Even the overall Wilson plot B is not very well determined, so I
>     wouldnt
>     worry too much..
>
>     Eleanor
>
>     Jiamu Du wrote:
>     > Dear All:
>     > According to your suggestion, I have set the peptide's occupency to
>     > 0.5. Two strategies were employed.
>     > 1. Direct using Refmac restrained refinement for 10 cycles. The B
>     > factor only drops to around 100. R/Rf did not change, either.
>     > 2. Direct CNS B-fator refinemen. The B factor drops to a moderate
>     > level 60-80, and the R/Rf each increases about 2%.
>     > 3. First using CNS B-fator refinemen nad next Refmac restrained
>     > refinement. The B factor drops to 60-80, and the R/Rf did not
>     change.
>     >
>     > I think next step TLS refinement should be carried out.
>     >
>     >
>     > On 4/30/07, *Philippe DUMAS* < [log in to unmask]
>     <mailto:[log in to unmask]>
>     > <mailto:[log in to unmask]
>     <mailto:[log in to unmask]>>> wrote:
>     >
>     >     Jiamu
>     >
>     >     According to the numbers you have mentioned I conclude that you
>     >     peptide occupancy should be around  60-64 %
>     >     I am interested to know what will be the value that you will
>     >     obtain after refinement...
>     >
>     >
>     >     Philippe Dumas
>     >     IBMC-CNRS, UPR9002
>     >     15, rue René Descartes 67084 Strasbourg cedex
>     >     tel: +33 (0)3 88 41 70 02
>     >     [log in to unmask] <mailto:[log in to unmask]>
>     <mailto: [log in to unmask] <mailto:[log in to unmask]>>
>     >
>     >
>     >         -----Message d'origine-----
>     >         *De :* CCP4 bulletin board [mailto:
>     [log in to unmask] <mailto:[log in to unmask]>
>     >         <mailto: [log in to unmask]
>     <mailto:[log in to unmask]>>]*De la part de* Jiamu Du
>     >         *Envoyé :* lundi 30 avril 2007 05:57
>     >         *À :* [log in to unmask]
>     <mailto:[log in to unmask]> <mailto:[log in to unmask]
>     <mailto:[log in to unmask]>>
>     >         *Objet :* [ccp4bb] extra high B factor
>     >
>     >         Dear All:
>     >         I am refining a protein-peptide complex struture at 2.6
>     >         angstrom resolution.
>     >         The data was obtain from a co-crystal and the wilson B
>     factor
>     >         of the data is about 70.
>     >         The affinity between protein and peptide is about 10E-7 to
>     >         10E-8 molar.
>     >         Protein fragment of the structure has a common B facor
>     about 50.
>     >         But surprisingly, the average B factor of the peptide is as
>     >         high as 130, although the peptide can be clearly traced
>     from
>     >         the the electron density map. All residues of the
>     peptide have
>     >         such a high B factor.
>     >         My question is how can I reduce the abnormal high B
>     factor to
>     >         a common level or if this high B factor acceptable.
>     >         And another question is if this high B fator will influence
>     >         the final refiment level.
>     >
>     >         Thanks.
>     >
>     >         --
>     >         Jiamu Du
>     >         State Key Laboratory of Molecular Biology
>     >         Institute of Biochemistry and Cell Biology Shanghai
>     Institutes
>     >         for Biological Sciences
>     >         Chinese Academy of Sciences (CAS)
>     >
>     >
>     >
>     >
>     > --
>     > Jiamu Du
>     > State Key Laboratory of Molecular Biology
>     > Institute of Biochemistry and Cell Biology Shanghai Institutes for
>     > Biological Sciences
>     > Chinese Academy of Sciences (CAS)
>
>
>
>
> -- 
> Jiamu Du
> State Key Laboratory of Molecular Biology
> Institute of Biochemistry and Cell Biology Shanghai Institutes for 
> Biological Sciences
> Chinese Academy of Sciences (CAS) 


-- 
Jeroen Raymundus Mesters, Ph.D.
Institut fuer Biochemie, Universitaet zu Luebeck
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