Re: [ccp4bb] How to reduce no. of overlaps

[Zhijie Li]

Hampton sells an adjustable mounted loop for this purpose.

 http://hamptonresearch.com/product_detail.aspx?cid=24&sid=136&pid=385


From: Frank von Delft 
Sent: Wednesday, March 07, 2012 1:26 AM
To: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] How to reduce no. of overlaps


You probably have to tilt your crystal, so that the long axis is parallel to 
the beam.  We do this routinely:  cut a plastic pipette tip to have a sharp 
point, then push the loop where it attaches to the pin, to bend the crystal 
itself.

You have to identify from your diffraction whether the long axis is pointing 
through the face or the edge of the loop.  As it's P6, chances are it's through 
the face, because long-axis P6 tends to make flat hexagons which lie flush with 
the face.  So you have to bend so the face of the loop upwards.

You'll have to practice this first, though, so put up an empty loop.  Top tips: 
* Don't breathe!  You'll blow the cryostream away.
* Bend the loop towards (rather than away from) the rim edge of the pin to 
which it's glued.
* Don't breathe!
* Practise practise practise.


Another thing:  most in-house sources allow you to reduce divergence of the 
beam.  You lose intensity, but no matter, just expose longer.  That also 
improves overlap.

Cheers
phx



On 07/03/2012 04:56, Dipankar Manna wrote: 
  Dear Crystallographers,



  I am working on a protein having SG P6, the cell parameters are a= 79, b= 79, 
c= 325. The crystals are forming in big size and with very good shape. It also 
diffracting very well in Home source facility both in terms of resolution and 
intensity. But the only problem is the number of overlaps. Its showing much 
more than the good spots. As a result the completeness is showing maximum up to 
65% even after collecting 180 degrees. I am unable to get a complete data. I 
tried with reducing the oscillation angel to 0.3 degree/0.5 degree but it did 
not improve that much. Please give me some suggestions.



  Regards,



  Dipankar





  Dipankar Manna

  Aurigene Discovery Technologies Limited,

  #39-40 KIADB Industrial Area, Electronic City, Phase II,

  Hosur Road, Bangalore- 560 100, India

  Cell: +91-9538631469  | Office Ph  : +91 80-66204422 (Extn: 398)  | Email ID: 
dipanka...@aurigene.com







--

  This e-mail and any files transmitted with it are for the sole use of the 
intended recipient(s) and may contain confidential and privileged 
information.If you are not the intended recipient, please contact the sender by 
reply e-mail and destroy all copies of the original message.Any unauthorized 
review, use, disclosure, dissemination, forwarding,printing or copying of this 
email or any action taken in reliance on this e-mail is strictly prohibited and 
may be unlawful.

  Visit us at http://www.aurigene.com

Re: [ccp4bb] How to reduce no. of overlaps

[Frank von Delft]

Yes... quite.  Thanks!

(Beam, rotation axis... how can you expect me to keep all these 
new-fangled inventions apart?!)




On 07/03/2012 07:33, VAN RAAIJ , MARK JOHAN wrote:

typo correction, you'll want the long axis parallel to the rotation
axis, not to the beam.
Mark

Quoting Frank von Delft:


You probably have to tilt your crystal, so that the long axis is
parallel to the beam.  We do this routinely:  cut a plastic pipette
tip to have a sharp point, then push the loop where it attaches to
the pin, to bend the crystal itself.

You have to identify from your diffraction whether the long axis is
pointing through the face or the edge of the loop.  As it's P6,
chances are it's through the face, because long-axis P6 tends to
make flat hexagons which lie flush with the face.  So you have to
bend so the face of the loop upwards.

You'll have to practice this first, though, so put up an empty loop.
  Top tips:
 * Don't breathe!  You'll blow the cryostream away.
 * Bend the loop towards (rather than away from) the rim edge of
the pin to which it's glued.
 * Don't breathe!
 * Practise practise practise.


Another thing:  most in-house sources allow you to reduce divergence
of the beam.  You lose intensity, but no matter, just expose longer.
  That also improves overlap.

Cheers
phx



On 07/03/2012 04:56, Dipankar Manna wrote:

Dear Crystallographers,

I am working on a protein having SG P6, the cell parameters are a=
79, b= 79, c= 325. The crystals are forming in big size and with
very good shape. It also diffracting very well in Home source
facility both in terms of resolution and intensity. But the only
problem is the number of overlaps. Its showing much more than the
good spots. As a result the completeness is showing maximum up to
65% even after collecting 180 degrees. I am unable to get a
complete data. I tried with reducing the oscillation angel to 0.3
degree/0.5 degree but it did not improve that much. Please give me
some suggestions.

Regards,

Dipankar

/Dipankar Manna/

*/Aurigene Discovery Technologies Limited,/*

/#39-40 KIADB Industrial Area, Electronic City, Phase II,/

/Hosur Road, Bangalore- 560 100, India/

/Cell: +91-9538631469 // | Office Ph  : +91 80-66204422 (Extn: 398)
  | Email ID: dipanka...@aurigene.com/




This e-mail and any files transmitted with it are for the sole use
of the intended recipient(s) and may contain confidential and
privileged information.If you are not the intended recipient,
please contact the sender by reply e-mail and destroy all copies of
the original message.Any unauthorized review, use, disclosure,
dissemination, forwarding,printing or copying of this email or any
action taken in reliance on this e-mail is strictly prohibited and
may be unlawful.

Visit us at http://www.aurigene.com



Mark J van Raaij
Laboratorio M-4
Dpto de Estructura de Macromoléculas
Centro Nacional de Biotecnología - CSIC
c/Darwin 3, Campus Cantoblanco
28049 Madrid
tel. 91 585 4616
email: mjvanra...@cnb.csic.es

Re: [ccp4bb] How to reduce no. of overlaps

[VAN RAAIJ , MARK JOHAN]

typo correction, you'll want the long axis parallel to the rotation  
axis, not to the beam.

Mark

Quoting Frank von Delft:

You probably have to tilt your crystal, so that the long axis is  
parallel to the beam.  We do this routinely:  cut a plastic pipette  
tip to have a sharp point, then push the loop where it attaches to  
the pin, to bend the crystal itself.


You have to identify from your diffraction whether the long axis is  
pointing through the face or the edge of the loop.  As it's P6,  
chances are it's through the face, because long-axis P6 tends to  
make flat hexagons which lie flush with the face.  So you have to  
bend so the face of the loop upwards.


You'll have to practice this first, though, so put up an empty loop.  
 Top tips:

* Don't breathe!  You'll blow the cryostream away.
* Bend the loop towards (rather than away from) the rim edge of  
the pin to which it's glued.

* Don't breathe!
* Practise practise practise.


Another thing:  most in-house sources allow you to reduce divergence  
of the beam.  You lose intensity, but no matter, just expose longer.  
 That also improves overlap.


Cheers
phx



On 07/03/2012 04:56, Dipankar Manna wrote:


Dear Crystallographers,

I am working on a protein having SG P6, the cell parameters are a=  
79, b= 79, c= 325. The crystals are forming in big size and with  
very good shape. It also diffracting very well in Home source  
facility both in terms of resolution and intensity. But the only  
problem is the number of overlaps. Its showing much more than the  
good spots. As a result the completeness is showing maximum up to  
65% even after collecting 180 degrees. I am unable to get a  
complete data. I tried with reducing the oscillation angel to 0.3  
degree/0.5 degree but it did not improve that much. Please give me  
some suggestions.


Regards,

Dipankar

/Dipankar Manna/

*/Aurigene Discovery Technologies Limited,/*

/#39-40 KIADB Industrial Area, Electronic City, Phase II,/

/Hosur Road, Bangalore- 560 100, India/

/Cell: +91-9538631469 // | Office Ph  : +91 80-66204422 (Extn: 398)  
 | Email ID: dipanka...@aurigene.com/





This e-mail and any files transmitted with it are for the sole use  
of the intended recipient(s) and may contain confidential and  
privileged information.If you are not the intended recipient,  
please contact the sender by reply e-mail and destroy all copies of  
the original message.Any unauthorized review, use, disclosure,  
dissemination, forwarding,printing or copying of this email or any  
action taken in reliance on this e-mail is strictly prohibited and  
may be unlawful.


Visit us at http://www.aurigene.com






Mark J van Raaij
Laboratorio M-4
Dpto de Estructura de Macromoléculas
Centro Nacional de Biotecnología - CSIC
c/Darwin 3, Campus Cantoblanco
28049 Madrid
tel. 91 585 4616
email: mjvanra...@cnb.csic.es

Re: [ccp4bb] How to reduce no. of overlaps

[Frank von Delft]

You probably have to tilt your crystal, so that the long axis is 
parallel to the beam.  We do this routinely:  cut a plastic pipette tip 
to have a sharp point, then push the loop where it attaches to the pin, 
to bend the crystal itself.


You have to identify from your diffraction whether the long axis is 
pointing through the face or the edge of the loop.  As it's P6, chances 
are it's through the face, because long-axis P6 tends to make flat 
hexagons which lie flush with the face.  So you have to bend so the face 
of the loop upwards.


You'll have to practice this first, though, so put up an empty loop.  
Top tips:

* Don't breathe!  You'll blow the cryostream away.
* Bend the loop towards (rather than away from) the rim edge of the 
pin to which it's glued.

* Don't breathe!
* Practise practise practise.


Another thing:  most in-house sources allow you to reduce divergence of 
the beam.  You lose intensity, but no matter, just expose longer.  That 
also improves overlap.


Cheers
phx



On 07/03/2012 04:56, Dipankar Manna wrote:


Dear Crystallographers,

I am working on a protein having SG P6, the cell parameters are a= 79, 
b= 79, c= 325. The crystals are forming in big size and with very good 
shape. It also diffracting very well in Home source facility both in 
terms of resolution and intensity. But the only problem is the number 
of overlaps. Its showing much more than the good spots. As a result 
the completeness is showing maximum up to 65% even after collecting 
180 degrees. I am unable to get a complete data. I tried with reducing 
the oscillation angel to 0.3 degree/0.5 degree but it did not improve 
that much. Please give me some suggestions.


Regards,

Dipankar

/Dipankar Manna/

*/Aurigene Discovery Technologies Limited,/*

/#39-40 KIADB Industrial Area, Electronic City, Phase II,/

/Hosur Road, Bangalore- 560 100, India/

/Cell: +91-9538631469 // | Office Ph  : +91 80-66204422 (Extn: 398)  | 
Email ID: dipanka...@aurigene.com/





This e-mail and any files transmitted with it are for the sole use of 
the intended recipient(s) and may contain confidential and privileged 
information.If you are not the intended recipient, please contact the 
sender by reply e-mail and destroy all copies of the original 
message.Any unauthorized review, use, disclosure, dissemination, 
forwarding,printing or copying of this email or any action taken in 
reliance on this e-mail is strictly prohibited and may be unlawful.


Visit us at http://www.aurigene.com

[ccp4bb] How to reduce no. of overlaps

[Dipankar Manna]

Dear Crystallographers,

I am working on a protein having SG P6, the cell parameters are a= 79, b= 79, 
c= 325. The crystals are forming in big size and with very good shape. It also 
diffracting very well in Home source facility both in terms of resolution and 
intensity. But the only problem is the number of overlaps. Its showing much 
more than the good spots. As a result the completeness is showing maximum up to 
65% even after collecting 180 degrees. I am unable to get a complete data. I 
tried with reducing the oscillation angel to 0.3 degree/0.5 degree but it did 
not improve that much. Please give me some suggestions.

Regards,

Dipankar


Dipankar Manna
Aurigene Discovery Technologies Limited,
#39-40 KIADB Industrial Area, Electronic City, Phase II,
Hosur Road, Bangalore- 560 100, India
Cell: +91-9538631469  | Office Ph  : +91 80-66204422 (Extn: 398)  | Email ID: 
dipanka...@aurigene.com





This e-mail and any files transmitted with it are for the sole use of the 
intended recipient(s) and may contain confidential and privileged 
information.If you are not the intended recipient, please contact the sender by 
reply e-mail and destroy all copies of the original message.Any unauthorized 
review, use, disclosure, dissemination, forwarding,printing or copying of this 
email or any action taken in reliance on this e-mail is strictly prohibited and 
may be unlawful.

Visit us at http://www.aurigene.com

Re: [ccp4bb] Zero occupancy and bad geometry

[Eleanor Dodson]

Coordinates with occ > 0 are refined and may shift - cds with occ=0 stay 
where they were.. This may result in bad geometry of course. You can 
"reglise" the whole residue in coot but that doesnt do nything except make 
it look tidier.. Eleanor


On Mar 7 2012, Joel Tyndall wrote:


Hi folks,

I have a case where I have changed the occupancy of a lysine residue to 
0.00 and after refinement the geometry is wrong, i.e. one of the bonds is 
extended (no connectivity in Coot and length is 1.76A). I have seen this 
before with other residues and a previous data set. The break in this 
case occurs between CG (1.00 occ) and CD (0.00 occ).


Any thoughts on why this is occurring?

Cheers

Joel

_
Joel Tyndall, PhD

Senior Lecturer in Medicinal Chemistry
National School of Pharmacy
University of Otago
PO Box 56 Dunedin 9054
New Zealand
Skype: jtyndall
http://www.researcherid.com/rid/C-2803-2008
Pukeka Matua
Te Kura Taiwhanga Putaiao
Te Whare Wananga o Otago
Pouaka Poutapeta 56 Otepoti 9054
Aotearoa

Ph / Waea   +64 3 4797293
Fax / Waeawhakaahua +64 3 4797034




--
Professor Eleanor Dodson
YSNL, Dept of Chemistry
University of York
Heslington YO10 5YW
tel: 00 44 1904 328259
Fax: 00 44 1904 328266

Re: [ccp4bb] qualification to serve as the co-author for the protein 3D structure stored in RCSB

[Eleanor Dodson]

Having done some of the work - written the paper - supervised the student 
who did a) and/or b)- etc etc.. Eleanor On Mar 7 2012, Dialing Pretty 
wrote:


Dear All,   Will you please show your opinion on the standards or 
qualifications to serve as the co-author for a protein 3D structure 
stored in RCSB?   Cheers,   Dialing  


--
Professor Eleanor Dodson
YSNL, Dept of Chemistry
University of York
Heslington YO10 5YW
tel: 00 44 1904 328259
Fax: 00 44 1904 328266

[ccp4bb] Postdoctoral position, microRNA biogenesis, Harvard Medical School

[Piotrek Sliz]

*Department: Biological Chemistry and Molecular Pharmacology
Location: Longwood Ave., Boston, MA
URL: 
http://hkl.hms.harvard.edu
Start Date: immediate/negotiable
Description: Our laboratory at Harvard Medical School has an immediate
opening for an protein/RNA crystallographer. Our group recently determined
high resolution X-ray structures of Lin28 with various members of let-7
microRNAs (Nam et al., Cell 2011), a system that has been implicated in
control of cell differentiation, cancer, and cell stemness. Projects are
available in our laboratory to build up on those recent results. As a
member of our group you will utilize a variety of structural biology
techniques, including X-ray Crystallography, SAXS, molecular dynamics and
NMR (in collaboration with Prof. James Chou). You might also collaborate
with members of Children’s Hospital Stem Cell Institute to perform
complementary biological experiments.

Candidates must hold PhD in biochemistry or related fields. Strong
background in molecular biology, biochemistry, large-scale protein
expression and purification of proteins and nucleic acids is desired.

Please submit: To apply, please send a single PDF file that incorporates CV
including contact information of 3 references and list of your experimental
expertise.

Person to contact: Dr. Piotrek Sliz
Surface mail address: Harvard Medical School. Department of Biological
Chemistry and Molecular Pharmacology. Center for Molecular and Cellular
Dynamics. 250 Longwood Ave., Rm SGM-130, Boston, MA, 02115.
Email address: ap...@hkl.hms.harvard.edu*

-- 


Piotrek Sliz
Assistant Professor in Pediatrics
Lecturer on BCMP

Harvard Medical School
Center for Molecular and Cellular Dynamics
http://hkl.hms.harvard.edu
Voice:  (617) 299-1455

[ccp4bb] Zero occupancy and bad geometry

[Joel Tyndall]

Hi folks,

I have a case where I have changed the occupancy of a lysine residue to 0.00 
and after refinement the geometry is wrong, i.e. one of the bonds is extended 
(no connectivity in Coot and length is 1.76A). I have seen this before with 
other residues and a previous data set. The break in this case occurs between 
CG (1.00 occ) and CD (0.00 occ).

Any thoughts on why this is occurring?

Cheers

Joel

_
Joel Tyndall, PhD

Senior Lecturer in Medicinal Chemistry
National School of Pharmacy
University of Otago
PO Box 56 Dunedin 9054
New Zealand
Skype: jtyndall
http://www.researcherid.com/rid/C-2803-2008
Pukeka Matua
Te Kura Taiwhanga Putaiao
Te Whare Wananga o Otago
Pouaka Poutapeta 56 Otepoti 9054
Aotearoa

Ph / Waea   +64 3 4797293
Fax / Waeawhakaahua +64 3 4797034

Re: [ccp4bb] qualification to serve as the co-author for the protein 3D structure stored in RCSB

[Bosch, Juergen]

Why don't you make a Venn diagram of Authors Deposition versus Authors 
Publication. The PDB provides the tools to download a spreadsheet with the 
necessary information.
Jürgen

On Mar 6, 2012, at 7:21 PM, Dialing Pretty wrote:

Dear All,

Will you please show your opinion on the standards or qualifications to serve 
as the co-author for a protein 3D structure stored in RCSB?

Cheers,

Dialing


..
Jürgen Bosch
Johns Hopkins University
Bloomberg School of Public Health
Department of Biochemistry & Molecular Biology
Johns Hopkins Malaria Research Institute
615 North Wolfe Street, W8708
Baltimore, MD 21205
Office: +1-410-614-4742
Lab:  +1-410-614-4894
Fax:  +1-410-955-2926
http://web.mac.com/bosch_lab/

[ccp4bb] qualification to serve as the co-author for the protein 3D structure stored in RCSB

[Dialing Pretty]

Dear All,
 
Will you please show your opinion on the standards or qualifications to serve 
as the co-author for a protein 3D structure stored in RCSB?
 
Cheers,
 
Dialing   

Re: [ccp4bb] Using intrinsically bound Zn atoms for phasing

[Pete Meyer]

Hi,
1. You don't mention how many Zn sites you have, or how big your protein 
is - as Phil mentioned, these are factors.
2. I'll add to the chorus - pick your wavelength(s) based on a 
fluorescence scan.
3. If "1.5A0" is your wavelength, not your resolution, you may still 
have some anomalous signal - I've had a dataset collected on a copper 
rotating anode (wavelength 1.54) with detectable zinc anomalous signal. 
 You'll get better signal closer to the anomalous peak; but if your 
Zn/protein ratio is large enough it may not be necessary.


Pete

Deepthi wrote:

Hi

I am trying to solve the structure of an engineered protein.The protein is 
crystallized with Zn bound to it  .We collected a 1.5A0 data. Molecular 
Replacement didn't yield a good match for the protein. I want to try MAD taking 
advantage of the Zn atoms in protein. I am not sure what wavelength should i 
use to collect the diffraction data for Zn. any suggestions?

Thank You
Deepthi

--
Deepthi

Re: [ccp4bb] Using intrinsically bound Zn atoms for phasing

[Eleanor Dodson]

For those who want to stay with the CCP4 GUI, yu can rn SHELXC/D/E from it 
and get the advantage of keeping all info in the same place.. The Zn edge 
is _ 1.28 but you can use the Zn signal for any wavelength less than that.. 
Eleanor


On Mar 6 2012, Bosch, Juergen wrote:

Since you've collected the data already use your favourite data 
processing program and treat the Friedel pairs separately. I'd suggest to 
try HKL2map in conjunction with SHELX C/D/E (sorry for the non CCP4 
advertisement here) for solving the heavy atom sites. You can in parallel 
also try SnB or BnP to find a substructure solution. Depending how bad 
you resulting density looks like you might want to improve your phases 
via Sharp.


If you want to stay in the CCP4 protected sandbox, then give Crank a try.

Jürgen

On Mar 6, 2012, at 3:24 PM, Francis E Reyes wrote:

http://skuld.bmsc.washington.edu/scatter/AS_form.html

Maybe useful to you.

However, I would advise to do a fluorescence scan over the range given in 
the graph and then use chooch to provide the precise energies for your 
peak and inflection.


If you have a large crystal don't expose all of it when you do the 
fluorescence scan but rather reserve a 'fresh' piece to do your actual 
data collection.


F


On Mar 6, 2012, at 1:09 PM, Deepthi wrote:

Hi

I am trying to solve the structure of an engineered protein.The protein 
is crystallized with Zn bound to it .We collected a 1.5A0 data. Molecular 
Replacement didn't yield a good match for the protein. I want to try MAD 
taking advantage of the Zn atoms in protein. I am not sure what 
wavelength should i use to collect the diffraction data for Zn. any 
suggestions?


Thank You
Deepthi

--
Deepthi

..
Jürgen Bosch
Johns Hopkins University
Bloomberg School of Public Health
Department of Biochemistry & Molecular Biology
Johns Hopkins Malaria Research Institute
615 North Wolfe Street, W8708
Baltimore, MD 21205
Office: +1-410-614-4742
Lab:  +1-410-614-4894
Fax:  +1-410-955-2926
http://web.mac.com/bosch_lab/







--
Professor Eleanor Dodson
YSNL, Dept of Chemistry
University of York
Heslington YO10 5YW
tel: 00 44 1904 328259
Fax: 00 44 1904 328266

Re: [ccp4bb] Using intrinsically bound Zn atoms for phasing

[Phil Jeffrey]

Self-referentially:

I once used the structural Zn of p53 to do a Zn MAD structure of a 
p53:53BP1 complex at 2.5 Angstrom with one zinc per 450 residues.

Apparently using 1.283, 1.282 and 1.262 Angstroms (i.e. the Zinc edge).
http://genesdev.cshlp.org/content/16/5/583.long
But of course do your own fluorescence scan.  The advantage of 
structural metals is full occupancy and relatively lower B-factor.


That map was actually pretty good, and since it came out of MLPHARE I 
don't doubt modern programs like SHARP could make it quite a lot better.


Phil Jeffrey
Princeton

On 3/6/12 3:09 PM, Deepthi wrote:

Hi

I am trying to solve the structure of an engineered protein.The protein
is crystallized with Zn bound to it .We collected a 1.5A0 data.
Molecular Replacement didn't yield a good match for the protein. I want
to try MAD taking advantage of the Zn atoms in protein. I am not sure
what wavelength should i use to collect the diffraction data for Zn. any
suggestions?

Thank You
Deepthi

--
Deepthi

Re: [ccp4bb] Using intrinsically bound Zn atoms for phasing

[Philippe DUMAS]

A reference for a real MAD phasing with Zinc (worked very well):
Ennifar et al. MAD phasing replacing magnesium with zinc. Acta Cryst.  
(2001). D57, 330


Philippe Dumas

"Bosch, Juergen"  a écrit :

Since you've collected the data already use your favourite data   
processing program and treat the Friedel pairs separately. I'd   
suggest to try HKL2map in conjunction with SHELX C/D/E (sorry for   
the non CCP4 advertisement here) for solving the heavy atom sites.   
You can in parallel also try SnB or BnP to find a substructure   
solution. Depending how bad you resulting density looks like you   
might want to improve your phases via Sharp.


If you want to stay in the CCP4 protected sandbox, then give Crank a try.

Jürgen

On Mar 6, 2012, at 3:24 PM, Francis E Reyes wrote:

http://skuld.bmsc.washington.edu/scatter/AS_form.html

Maybe useful to you.

However, I would advise to do a fluorescence scan  over the range   
given in the graph and then use chooch to provide the precise   
energies for your peak and inflection.


If you have a large crystal don't expose all of it when you do the   
fluorescence scan but rather reserve a 'fresh' piece to do your   
actual data collection.


F


On Mar 6, 2012, at 1:09 PM, Deepthi wrote:

Hi

I am trying to solve the structure of an engineered protein.The   
protein is crystallized with Zn bound to it  .We collected a 1.5A0   
data. Molecular Replacement didn't yield a good match for the   
protein. I want to try MAD taking advantage of the Zn atoms in   
protein. I am not sure what wavelength should i use to collect the   
diffraction data for Zn. any suggestions?


Thank You
Deepthi

--
Deepthi

..
Jürgen Bosch
Johns Hopkins University
Bloomberg School of Public Health
Department of Biochemistry & Molecular Biology
Johns Hopkins Malaria Research Institute
615 North Wolfe Street, W8708
Baltimore, MD 21205
Office: +1-410-614-4742
Lab:  +1-410-614-4894
Fax:  +1-410-955-2926
http://web.mac.com/bosch_lab/

Re: [ccp4bb] Using intrinsically bound Zn atoms for phasing

[Bosch, Juergen]

Since you've collected the data already use your favourite data processing 
program and treat the Friedel pairs separately. I'd suggest to try HKL2map in 
conjunction with SHELX C/D/E (sorry for the non CCP4 advertisement here) for 
solving the heavy atom sites. You can in parallel also try SnB or BnP to find a 
substructure solution. Depending how bad you resulting density looks like you 
might want to improve your phases via Sharp.

If you want to stay in the CCP4 protected sandbox, then give Crank a try.

Jürgen

On Mar 6, 2012, at 3:24 PM, Francis E Reyes wrote:

http://skuld.bmsc.washington.edu/scatter/AS_form.html

Maybe useful to you.

However, I would advise to do a fluorescence scan  over the range given in the 
graph and then use chooch to provide the precise energies for your peak and 
inflection.

If you have a large crystal don't expose all of it when you do the fluorescence 
scan but rather reserve a 'fresh' piece to do your actual data collection.

F


On Mar 6, 2012, at 1:09 PM, Deepthi wrote:

Hi

I am trying to solve the structure of an engineered protein.The protein is 
crystallized with Zn bound to it  .We collected a 1.5A0 data. Molecular 
Replacement didn't yield a good match for the protein. I want to try MAD taking 
advantage of the Zn atoms in protein. I am not sure what wavelength should i 
use to collect the diffraction data for Zn. any suggestions?

Thank You
Deepthi

--
Deepthi

..
Jürgen Bosch
Johns Hopkins University
Bloomberg School of Public Health
Department of Biochemistry & Molecular Biology
Johns Hopkins Malaria Research Institute
615 North Wolfe Street, W8708
Baltimore, MD 21205
Office: +1-410-614-4742
Lab:  +1-410-614-4894
Fax:  +1-410-955-2926
http://web.mac.com/bosch_lab/

Re: [ccp4bb] Using intrinsically bound Zn atoms for phasing

[Francis E Reyes]

http://skuld.bmsc.washington.edu/scatter/AS_form.html

Maybe useful to you. 

However, I would advise to do a fluorescence scan  over the range given in the 
graph and then use chooch to provide the precise energies for your peak and 
inflection. 

If you have a large crystal don't expose all of it when you do the fluorescence 
scan but rather reserve a 'fresh' piece to do your actual data collection. 

F


On Mar 6, 2012, at 1:09 PM, Deepthi wrote:

> Hi
> 
> I am trying to solve the structure of an engineered protein.The protein is 
> crystallized with Zn bound to it  .We collected a 1.5A0 data. Molecular 
> Replacement didn't yield a good match for the protein. I want to try MAD 
> taking advantage of the Zn atoms in protein. I am not sure what wavelength 
> should i use to collect the diffraction data for Zn. any suggestions?
> 
> Thank You
> Deepthi
> 
> -- 
> Deepthi

[ccp4bb] Using intrinsically bound Zn atoms for phasing

[Deepthi]

Hi

I am trying to solve the structure of an engineered protein.The protein is
crystallized with Zn bound to it  .We collected a 1.5A0 data. Molecular
Replacement didn't yield a good match for the protein. I want to try MAD
taking advantage of the Zn atoms in protein. I am not sure
what wavelength should i use to collect the diffraction data for Zn. any
suggestions?

Thank You
Deepthi

-- 
Deepthi

[ccp4bb] Petition to Increase NIH Budget

[Shiva Bhowmik]

Dear CCP4-ers,

Sorry for the offline topic. I would like to bring this the following
petition:

Dear Colleagues,
I wanted to bring your attention to a petition (see link below) started at
WhiteHouse.gov by our colleague Dr. Steve Meltzer
at Johns Hopkins that calls for an increase for the NIH budget. As you
know, the President recently announced his proposed FY2013 budget, which
called for a flat NIH budget. After factoring in inflation, flat funding is
an equivalent of a funding cut. Research by biomedical community is already
heavily stressed, with NIH funding rates at a record low. Many of you would
be interested in supporting this petition, which provides a unique
opportunity to gain the attention of the White House. If the petition
receives 25,000 signatures by March 18th, the White House will review it.
We currently have over 12,000 signatures.

These are troubling times for the research community and medical progress
is at great risk.  Investigators, their institutions, Private research
funding organizations, patient advocate groups, and the biotech/pharma
sector all have a common goal - the betterment of human health. NIH funding
is a critical component for the biomedical research pipeline and increasing
funding for the NIH should be a goal we can all work together on. Please
help us in this endeavor and sign and share this petition with your
friends, family, and colleagues.

The petition link:

http://wh.gov/81O

best regards,
Kerri

Kerri A. Mowen, Ph.D.
Assistant Professor
Departments of Chemical Physiology & Immunology and Microbial Sciences The
Scripps Research Institute 10550 N. Torrey Pines Rd, IMM-11 Immunology
Building Room 315 La Jolla, CA 92037
tel: 858-784-2248

Support bettering human health and improving our economy! Sign this
petition to support  increasing funding for the National Institutes of
Health:
http://wh.gov/81O

Re: [ccp4bb] HKL2000 indexing problem

[Nic Steussy]

  
  
Our most common problem is
  an incorrect beam center.  The crystal we collect most frequently
  has large cell dimensions, so if the beam center is off even a bit
  it will bollix up the indexing.  What we usually do at the
  synchrotron is look for a refined beam center from the group there
  before us.  That has been our secret to success on a number of
  occasions.
  
  Nic out
  

On 02/20/2012 07:28 PM, Peter Hsu wrote:

  Hi all,

I recently collected a data set off a single crystal and have had problems with indexing it. Every time I go pick peaks for indexing it constantly picks peaks that are just slightly off the actual peak. After indexing, it would always be that 2 of the 3 cell dimensions would be fairly normal, while the 3rd would have some impossible value such as 1. 

On some other occasions if it manages to pick peaks properly, and every time I go to index it, it gives back an error that I don't have enough peaks picked to index (picked nearly 500).

I've tried using a number of different images to index from and have run into the same problem.

Has anyone else run into these problems? Does anyone have any idea what might be wrong w/my dataset and/or crystal?

Thanks in advance for any insight,

Peter



-- 

C. Nicklaus Steussy, M.D., Ph.D.
Purdue University
csteu...@purdue.edu



  

Re: [ccp4bb] stereo

[Nic Steussy]

  
  
I have a Quadro 3800
  paired with an Acer GD235Hz with the computer running Ubuntu 10.04
  LTS and the standard NVIDIA linux driver.  Hardware stereo works
  pretty well in Coot, though it has a habit of inverting
  occasionally.  Pymol works well.  Have not tried O.
  
  Nic out
  

On 02/23/2012 02:16 PM, Hena Dutta wrote:
Hello,
  Is anyone using nvidia quadro 4000 graphics card for linux
  environment to see stereo in pymol, coot, O or similar programs?
  If yes, I like to buy that.
  Thanks...
  Hena


-- 

C. Nicklaus Steussy, M.D., Ph.D.
Purdue University
csteu...@purdue.edu



  

[ccp4bb] off-topic micro ProDiCon

[Emily]

Hi all, 

I am in search of an old device called the micro ProDiCon.  It seems to be 
discontinued commercially but I thought there might be some hoarders out there 
that might be willing to give theirs up for a fee.  Please e mail me if you 
have any leads.

Thanks,
Emily

[ccp4bb] job vacancy at Laval University

[Clément Morin]

Structural biologists targeting breast cancer at Laval University Medical 
Center (CHUL, CHUQ)

Postdoctoral and Ph.D. positions are available at Laval University Hospital 
Research Center (CHUL, CHUQ), for the study of biological function of important 
enzymes and receptors implied in breast cancer development using structural 
biology, cell biology, genomics and proteomics. Our research involves related 
collaborative projects in steroid enzyme structure-function and in estrogen 
synthesis pathways, supported by the Canadian Institutes of Health Research 
(CIHR). Inhibitors of steroid enzymes will be searched for breast cancer 
treatment and will be tested in cancer cells and animal models. CIHR supported 
collaborative projects for anti-viral drugs are also available in the 
laboratory.

Since our determination of the first human steroid enzyme structure, the type 1 
17beta-hydroxysteroid dehydrogenase, we have analyzed a series of protein 
structures, resulting in leads for the cancer therapy. Expertise in one or two 
domains in crystallography, enzymology, cell biology, genomics and proteomics 
are highly welcome. Quebec City, with its original European style, is alongside 
the St. Laurence River, providing a beautiful and an excellent research 
environment. All positions will be well remunerated at CIHR levels, while the 
living expenses are modest in Québec City. 

For those who are interested in the above projects, please send your C.V. to 
sx...@crchul.ulaval.ca

Re: [ccp4bb] REFMAC Riding Hydrogens

[Ian Tickle]

> Pavel’s statement is likely a bit of an exaggeration, but he has a valid
> (yet hard to prove point). The default in CCP4i was (and is?) to use
> hydrogens only if present in the input file. This is IMO not a safe default.

I agree: what I find confusing here is that the current CCP4i default
is indeed to use H atoms only if present in input, whereas the current
documentation 
(http://www.ccp4.ac.uk/html/refmac5/keywords/restraints.html#makecif)
says (quoting verbatim):

HYDR [ Yes | No | All ] (Default HYDRogens All)
How to deal with the hydrogen atoms.
Y - if hydrogens are present in the input file, use them,
N - ignore hydrogens even if they are present in the input coordinate file,
A - add all hydrogens in their riding positions and use them for
geometry and structure factor calculations. They also contribute to
the gradient and second derivative matrix of geometry residual.

The documentation in this case also corresponds to the source code default:

  MAKE_HFLAG = 'A'

Which do we believe: GUI or source code?  Personally I'm much more
inclined to trust the defaults in the code over either the
documentation or the GUI, because I have a strong suspicion that the
CCP4i defaults (and yes even on occasions the documentation!) lag
somewhat behind the "community view" on this as represented by the
current code version.  Or is this an incorrect perception?

> Because there were some reporting errors in the past
> (http://proteincrystallography.org/ccp4bb/message18808.html) it is hard to
> tell from the PDB when refinement with hydrogens became hip. Discussions on
> this BB show that at the use of riding hydrogens is still not fully
> accepted, especially at low resolution (where they actually help most).

Documentation says: "For low resolution and early stages it is better
to use MAKE HYDR N, i.e. do not use hydrogens even if they are present
in the input file.".

Cheers

-- Ian

Re: [ccp4bb] REFMAC Riding Hydrogens

[Robbie Joosten]

Hi Everyone, 

 

Pavel’s statement is likely a bit of an exaggeration, but he has a valid (yet 
hard to prove point). The default in CCP4i was (and is?) to use hydrogens only 
if present in the input file. This is IMO not a safe default. 

Because there were some reporting errors in the past 
(http://proteincrystallography.org/ccp4bb/message18808.html) it is hard to tell 
from the PDB when refinement with hydrogens became hip. Discussions on this BB 
show that at the use of riding hydrogens is still not fully accepted, 
especially at low resolution (where they actually help most). 

 

Cheers,

Robbie

 

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Pavel 
Afonine
Sent: Monday, March 05, 2012 21:53
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] REFMAC Riding Hydrogens

 

Dear Tim,

 

good catch, thanks; I could craft that phrase more carefully! Although often it 
may not be quite fair to take phrases out of context: this newsletter article 
was written in the context of macromolecular refinement. And yes, "recently" 
may be a broad term -:)

 

All the best,

Pavel

On Mon, Mar 5, 2012 at 12:45 PM, Tim Gruene  wrote:

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Hash: SHA1

Dear Pavel,

you may want to add to the structures mentioned in [1] one or two
organic structures present in the Cambridge Database.

"Until recently it was customary to ignore hydrogen atoms throughout the
process of crystallographic X-­‐ray structure determination." [1]

'recently' as in 1997 [2]? Even though 1997 is probably a poor
estimation of the corresponding year...

Cheers,
Tim


[1] "On contribution of hydrogen atoms to X-ray scattering"
http://www.phenix-online.org/newsletter/
[2] http://shelx.uni-ac.gwdg.de/SHELX/shelx.pdf


On 03/05/2012 09:14 PM, Pavel Afonine wrote:
> Hi,
>
> On Mon, Mar 5, 2012 at 11:52 AM, Matthew Franklin wrote:
>
>> Adding the riding hydrogens generally gives you some improvement in R
>> factors even with a good quality (i.e. stereochemically correct) model.
>>
>
> and here are the results of more or less systematic test that prove this:
>
> see "On contribution of hydrogen atoms to X-ray scattering"
> here:
> http://www.phenix-online.org/newsletter/
>
> Pavel
>

- --
Dr Tim Gruene
Institut fuer anorganische Chemie
Tammannstr. 4
D-37077 Goettingen

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