Re: [ccp4bb] problem with HKL2000 license

[Frank von Delft]

Make sure the command "hostname" returns the same value as it did before the
reinstall -- check in the cr_info file that access_prod produces. 
 
Almost always, it's that the domain name is either now added or
alternatively removed, i.e. that you used to have "machine.some.domain.edu",
and not it's just "machine" (or vice versa).
 
Hope that helps.
phx


  _  

From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of yang
li
Sent: 30 April 2007 04:49
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] problem with HKL2000 license


Hi,All,
Now I am installing HKL2000 and met a curious problem:
The license cr_info was apllied before, the system is fc6 at that time.
Before received the license,
I reinstalled the system for some reason, now the system is fc4. the install
files and anom.dat file
was copied from other people. I changed all the configs,now if I run
access_prod file the values of
HOST-NAME, HOST-ID,HOSTNAME,HW-PROV,CPU-SERIAL are all same with the values
when
apllied the license. But now if I type HKL2000, comes the error information:
ERROR: Not a valid HKL-2000 license: Problem with hardware recognition

Field HOST-NAME 

Error code: 6

 

I googled for this error, the answer is: HOST-NAME line in the info file
must have changed. Please re-run the access_prod program on that computer
and send to [EMAIL PROTECTED] Your access file, cr_info, needs to be
updated.

 

   I wonder if I rerun the access_prod programm, the hostname is the same,
how could it give this error? 

It is a little troublesome if have to update the license, so I think if
anyone can tell the reason, it will save much work. 

 

Best Regards

 

Re: [ccp4bb] NCS-averaged composite omit map?

[Eleanor Dodson]

Use COOT to do NCS averaging then look at the maps of B onto A a well as 
the averaged one to see where differences might be.

Eleanor

Paul Paukstelis wrote:
I'm working on a 4.5-A structure with 4-fold NCS. I've generated a SA 
composite omit map and all the protomers look pretty good, but with 
weaker density in mostly different parts for each. Am I correct that 
CNS doesn't do any NCS averaging automatically, and is there any 
reason why I shouldn't average the density with NCS matrix/protomer mask?


--paul

Re: [ccp4bb] buried surface area

[Eleanor Dodson]

I use PISA  - it tells you lots of things and I think will do this as well

http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html

You can download your coordinates.. Will need to name subunits A and B 
or something I think..


Eleanor


Gloria Borgstahl wrote:

Hello all,
What is the easiest way, these days, to calculate the buried surface area
between two subunits of a protein?
Thanks, and Happy Friday, Gloria
**
Gloria Borgstahl
Eppley Institute for Cancer Research and Allied Diseases
987696 Nebraska Medical Center   
10732A Lied Transplant Center   
Omaha, NE 68198-7696


http://sbl.unmc.edu
Office (402) 559-8578
FAX (402) 559-3739

Associate Professor
Interests:  Cancer, Biochemistry, DNA Metabolism,  Protein Crystallography, 
Modulated Crystals,  Crystal Perfection, X-ray Topography,

Varley, Rowling, Avatar, BRAN, RAGBRAI, swimming & soccer
**

[ccp4bb] Postdoc position in Korea (Samsung Biomedical Research Institute)

[Kyeong Kyu Kim]

Postdoctoral Position - Protein Crystallography

Samsung Biomedical Research Institute

Sungkyunkwan University School of Medicine

Suwon, Korea

 

A post-doctoral position in protein crystallography is immediately available
at Sungkyunkwan University School of Medicine/Samsung Biomedical Research
institute in Suwon, Korea. The major project aims to solve the crystal
structure of several hydrolases from Thermococcus sp. In this project,
purified proteins will be provided for structure-determination. The second
project will focus on proteins involved in protein quality control and
ubiquitin processing. This project will be complemented by functional
experiments using molecular biology and biochemistry, as well as alternative
biophysical techniques.  Hence, the applicant will be exposed to a variety
of disciplines.

 

The unit is newly equipped with automatic crystallization robot, X-ray
facility comprising a Rigaku rotating anode X-ray generator, R-Axis IV dual
imaging plate detector with confocal mirror optics, and X-stream
cryo-cooling system. Recent crystallographic studies in the lab also have
utilized synchrotron sources in Pohang Synchrotron Light Source (Korea), USA
(NSLS and ALS) and in Japan (spring-8 and Photon Factory). There are
excellent facilities for molecular biology, protein overexpression, and
biochemistry in the unit.

 

The applicant should have a PhD in crystallography, or a related research
area such as chemistry, biophysics, biochemistry or molecular biology.
However, experience in crystal structure determination is preferred.
Interested applicants please send a CV with the names of three references to
address below. E-mail inquiries about the position are welcome. Review of
applications begins May 1, 2007 and will continue until the position is
filled.

 

Kyeong Kyu Kim, Ph. D.

Department of Molecular Cell Biology

Samsung Biomedical Research Institute

Sungkyunkwan University School of Medicine

Suwon 440-746, Korea

tel : 82-31-299-6136

fax : 82-31-299-6159

e-mail :   [EMAIL PROTECTED]

web: http://smsb1.skku.ac.kr

 

 

Kyeong Kyu Kim

 

Re: [ccp4bb] weird thing about MR

[Eleanor Dodson]

You say you have only found one of at least 3 copies?
Why not continue and find the other 2 before starting refinement? Phaser 
will do that automatically if you ask for 3 molecules..

Refinement of structural fragments is always tricky..
Eleanor

Yi Xue wrote:

Dear all:

  I have a dataset diffracted to about 3A, and it is supposed to be
protein/DNA complex. The spacegroup could be I4, I422, I222, or F222.
  I used the protein structure to start molecular replacement. There are
three copies of complexes if assuming I422 symmetry. For the search of the
first copy (phaser),  the top solution had very good RFZ (7.4) and TFZ
(18.1), meanwhile, there were total 44 good solutions, with RFZ>7 and
TFZ>14.  However, subsequent rigid body refinment  of the top solution gave
very high R and Rfree of around 80%.  I tried all choices of space groups,
briefly, they behaved similarly: MR found good solutions, R and Rfree were
weirdly high.

 I was thinking maybe the spacegroup is wrong, I checked the data very
carefully, and it seemed that the spacegroups I had chosen should be right.
 Any comments are highly appreciated.


Best
Yi


  

[ccp4bb] TFT monitors n stereo

[Vineet Gaur]

Hi all
i have a non CCP4 query.
pls tell me how to view stereo images on a TFT monitor.

thanx

vineet gaur

Re: [ccp4bb] extra high B factor

[Jiamu Du]

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 <[EMAIL PROTECTED]> 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
[EMAIL PROTECTED]


-Message d'origine-
*De :* CCP4 bulletin board [mailto: [EMAIL PROTECTED] la part de*Jiamu Du
*Envoyé :* lundi 30 avril 2007 05:57
*À :* CCP4BB@JISCMAIL.AC.UK
*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)

Re: [ccp4bb] extra high B factor

[Eleanor Dodson]

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 

Occ = 0.5  

Occ = 0.33  

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* <[EMAIL PROTECTED] 
> 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
[EMAIL PROTECTED] 
 


-Message d'origine-
*De :* CCP4 bulletin board [mailto: CCP4BB@JISCMAIL.AC.UK
]*De la part de* Jiamu Du
*Envoyé :* lundi 30 avril 2007 05:57
*À :* CCP4BB@JISCMAIL.AC.UK 
*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) 

Re: [ccp4bb] extra high B factor

[Jiamu Du]

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 <[EMAIL PROTECTED]> 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 

Occ = 0.5  

Occ = 0.33  

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* <[EMAIL PROTECTED]
> > 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
> [EMAIL PROTECTED] 
>
>
> -Message d'origine-
> *De :* CCP4 bulletin board [mailto: CCP4BB@JISCMAIL.AC.UK
> ]*De la part de* Jiamu Du
> *Envoyé :* lundi 30 avril 2007 05:57
> *À :* CCP4BB@JISCMAIL.AC.UK 
> *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)

Re: [ccp4bb] extra high B factor

[artis]

Shelx can refine occupancies.
Arti
> 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 <[EMAIL PROTECTED]> 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 
>>
>> Occ = 0.5  
>>
>> Occ = 0.33  
>>
>> 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* <[EMAIL PROTECTED]
>> > > 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
>> > [EMAIL PROTECTED] 
>> >
>> >
>> > -Message d'origine-
>> > *De :* CCP4 bulletin board [mailto: CCP4BB@JISCMAIL.AC.UK
>> > ]*De la part de* Jiamu Du
>> > *Envoyé :* lundi 30 avril 2007 05:57
>> > *À :* CCP4BB@JISCMAIL.AC.UK 
>> > *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)
>


Arti S. Pandey
Graduate Student
Chemistry and Biochemistry
Montana State University
Bozeman,MT 59717

Re: [ccp4bb] extra high B factor

[Manfred S. Weiss]

Dear all,

Frankly speaking I am having some doubts about this whole
discussion.

1. Apparently, it does not make a difference in R and Rfree
   whether the peptide is in the structure or not. This does
   suggest that there is very little (if any) information about
   the peptide in the data. Right?

2. You stated that you can unambiguously trace the peptide
   from N- to C-terminus. If you assume atomic B-factors of
   100 or larger and calculate the density, I really can't
   see how this is possible. Maybe you could produce an
   omit S. A. difference electron density map to show this.

3. You said the affinity between protein and peptide is
   10^-7 or 10^-8 M. This means that something is a bit
   strange in any case. With this affinity you should get
   100% occupancy. Of course, it is possible that your
   buffer/cryo-solvent/etc. reduce the affinity. Have you
   considered this?

Cheers, Manfred.


*  *
*Dr. Manfred S. Weiss  *
*  *
* Team Leader  *
*  *
* EMBL Hamburg OutstationFon: +49-40-89902-170 *
* c/o DESY, Notkestr. 85 Fax: +49-40-89902-149 *
* D-22603 Hamburg   Email: [EMAIL PROTECTED] *
* GERMANY   Web: www.embl-hamburg.de/~msweiss/ *
*  *



On Mon, 30 Apr 2007, Jiamu Du wrote:

> 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 <[EMAIL PROTECTED]> 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 
> >
> > Occ = 0.5  
> >
> > Occ = 0.33  
> >
> > 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* <[EMAIL PROTECTED]
> > > > 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 Ren? Descartes 67084 Strasbourg cedex
> > > tel: +33 (0)3 88 41 70 02
> > > [EMAIL PROTECTED] 
> > >
> > >
> > > -Message d'origine-
> > > *De :* CCP4 bulletin board [mailto: CCP4BB@JISCMAIL.AC.UK
> > > ]*De la part de* Jiamu Du
> > > *Envoy? :* lundi 30 avril 2007 05:57
> > > *? :* CCP4BB@JISCMAIL.AC.UK 
> > > *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.
> > 

Re: [ccp4bb] extra high B factor

[mesters]

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* <[EMAIL PROTECTED] 
> 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 

Occ = 0.5  

Occ = 0.33  

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* < [EMAIL PROTECTED]

> >> 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
> [EMAIL PROTECTED] 
mailto:[EMAIL PROTECTED]>>
>
>
> -Message d'origine-
> *De :* CCP4 bulletin board [mailto:
CCP4BB@JISCMAIL.AC.UK 
> mailto:CCP4BB@JISCMAIL.AC.UK>>]*De la part de* Jiamu Du
> *Envoyé :* lundi 30 avril 2007 05:57
> *À :* CCP4BB@JISCMAIL.AC.UK
 >
> *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
Zentrum fuer Medizinische Struktur und Zellbiologie
Ratzeburger Allee 160, D-23538 Luebeck
Tel: +49-451-5004070, Fax: +49-451-5004068
E-mail: [EMAIL PROTECTED]
Http://www.biochem.uni-luebeck.de
Http://www.iobcr.org
Http://www.opticryst.org
--
If you can look into the seeds of time and say
which grain will grow and which will not - speak then to me  (Macbeth)
--

[ccp4bb] AW: [ccp4bb] extra high B factor

[Herman . Schreuder]

Dear All,

What I read from the email is that the R/Rfree values do not change much if you 
refine with high occupancy-high B or with low occupancy-low B, which is what 
one expects with 2.6 Å data. This is something different from leaving out the 
inhibitor from the refinement. 

Considering the affinity, one can not compare the affinity free in solution 
with the affinity in a crystal. Crystal packing may hinder peptide binding. 
Another, very important thing is to add sufficient peptide to the 
cryoprotectant, otherwise one may loose almost all bound peptide during the 
cryoprotection. What often works for us, is just trying another crystal, we 
sometimes find very different inhibitor-occupancies for (almost) identical 
conditions.

Best regards,
Herman


-Ursprüngliche Nachricht-
Von: CCP4 bulletin board [mailto:[EMAIL PROTECTED] Im Auftrag von Manfred S. 
Weiss
Gesendet: Montag, 30. April 2007 16:32
An: CCP4BB@JISCMAIL.AC.UK
Betreff: Re: [ccp4bb] extra high B factor

Dear all,

Frankly speaking I am having some doubts about this whole discussion.

1. Apparently, it does not make a difference in R and Rfree
   whether the peptide is in the structure or not. This does
   suggest that there is very little (if any) information about
   the peptide in the data. Right?

2. You stated that you can unambiguously trace the peptide
   from N- to C-terminus. If you assume atomic B-factors of
   100 or larger and calculate the density, I really can't
   see how this is possible. Maybe you could produce an
   omit S. A. difference electron density map to show this.

3. You said the affinity between protein and peptide is
   10^-7 or 10^-8 M. This means that something is a bit
   strange in any case. With this affinity you should get
   100% occupancy. Of course, it is possible that your
   buffer/cryo-solvent/etc. reduce the affinity. Have you
   considered this?

Cheers, Manfred.


*  *
*Dr. Manfred S. Weiss  *
*  *
* Team Leader  *
*  *
* EMBL Hamburg OutstationFon: +49-40-89902-170 *
* c/o DESY, Notkestr. 85 Fax: +49-40-89902-149 *
* D-22603 Hamburg   Email: [EMAIL PROTECTED] *
* GERMANY   Web: www.embl-hamburg.de/~msweiss/ *
*  *



On Mon, 30 Apr 2007, Jiamu Du wrote:

> 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 <[EMAIL PROTECTED]> 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 
> >
> > Occ = 0.5  
> >
> > Occ = 0.33  
> >
> > 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* <[EMAIL PROTECTED] 
> > > > 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
> > > [EMAIL PROTECTED] 
> > >
> > >
> > > -Message d'origine-
> > > *De :* CCP4 bulletin board [mailto: CCP4BB@JISCMAIL.AC.UK
> > > ]*De la part de* Jiamu Du
> > >

Re: [ccp4bb] extra high B factor

[Bernhard Rupp]

> If you assume atomic B-factors of 100 or larger and 
> calculate the density, I really can't see how this is possible. 

Well, I do. The recipe has been published in NSB as a cautionary tale.
http://www.ruppweb.org/cvs/br/rupp_2001_NSB_questions_BotA.pdf
and there is quite fail-safe away to investigate
http://www.ruppweb.org/Xray/tutorial/rscc.htm

BR

[ccp4bb] Electron Microscopist Position

[Tuli Mukhopadhyay]

Electron Microscopist Researcher and Facility Manager

Indiana University in Bloomington (IUB) has a position open for an 
Electron Microscopist Researcher and Facility Manager. IUB is purchasing 
a 300 kV FEG TEM designed for both biological and material science 
applications, including cryo, tomography, EDS, and EELS spectroscopy. 
Qualified applicants will have at least 2 years of experience, priority 
given to those with cryo, low-dose, or tomography experience. 
Responsibilities include data collection, user training, and scope 
maintenance.  Possibility for teaching lectures and workshops, if 
interested.  Start date: September 1.  Applications will be reviewed 
starting May 15 and until the position is filled.  Please send CV, a 
brief research statement including list of publications, EM 
techniques/applications, data processing and image reconstruction 
experience.  Please send all materials to:  Dr. Bogdan Dragnea, 
Department of Chemistry, Indiana University, 800 E. Kirkwood Ave., 
Bloomington, IN  47405, or electronically to [EMAIL PROTECTED] 
Indiana University is an Equal Opportunity Affirmative Action employer.

[ccp4bb] Buried Surface Area - summary

[Gloria Borgstahl]

Here was the query.What is the easiest way, these days, to calculate the buried surface area between two subunits of a protein? Here are the software and links received (thank you all):1. 4+ votes for PISA  - This following website is great and does a very nice job of analyzing protein interfaces (including, but not limited to, buried surface area calculations):http://www.ebi.ac.uk/msd-srv/prot_int/pistart.htmlFYI. the next website can also yield useful infoto figure out if an interface is physiologically relevant or not (less detailed output though):http://www.ebi.ac.uk/thornton-srv/databases/pita/2. surface ( http://www.ccp4.ac.uk/dist/html/surface.html ).   3. Two or three votes for areaimol in CCP4 works well - but beware that for cases we have worked on the default value for point density needs to be increased to get accurate values. With the subunits A and B and their surfaces surf(A) and surf(B) the equation should hold: surf(A) + surf(B) = surf(A+B) +2x where x is the surface between them. So if you calculate the surfaces of each subunit separately and of the complex that yields x. 4. NACCESS  http://wolf.bms.umist.ac.uk/naccess/ 5.  CCP4mg will do this - and show you graphically..http://www.ysbl.york.ac.uk/~ccp4mg/ccp4mg_help/analysis.html#sas6.  see the discussion and references in: http://xray.bmc.uu.se/cgi-bin/gerard/reprint_mailer.pl?pref=847.  Chimera.http://www.cgl.ucsf.edu/pipermail/chimera-users/2006-December/001131.htmlThanks again.**Gloria BorgstahlEppley Institute for Cancer Research and Allied Diseases987696 Nebraska Medical Center   10732A Lied Transplant Center   Omaha, NE 68198-7696http://sbl.unmc.eduOffice (402) 559-8578FAX (402) 559-3739Associate ProfessorInterests:  Cancer, Biochemistry, DNA Metabolism,  Protein Crystallography, Modulated Crystals,  Crystal Perfection, X-ray Topography,Varley, Rowling, Avatar, BRAN, RAGBRAI, swimming & soccer**