Re: [ccp4bb] Sad News

[James Holton]

Ward was the Program Official for all my grants!  My life will not be 
the same without him.  He was always so supportive and helpful with 
advice on how to navigate the sometimes convoluted system that is the 
NIH.  From the Protein Structure Initiative to today his hard work has 
made possible my entire scientific career.


"missed" doesn't seem to cover it. May your rest be a peaceful one, Ward.

-James Holton
MAD Scientist

On 7/18/2020 4:36 AM, Sweet, Robert wrote:

I'm writing to acknowledge the passing of Ward Smith during the weekend of 5 
July. Ward got his PhD with Martha Ludwig at U. of Michigan, and then came to 
UCLA in 1977 to join Dave Eisenberg’s group as a postdoc. During the course of 
things, he met Cheryl Janson, a Paul Boyer postdoc, and they were married in 
1980.  During his time at UCLA Ward became expert in operation of the 
Xuong/Hamlin/Nielsen multi-wire system at UCSD and tutored the UCLA users in 
its use. In 1985 Ward and Cheryl left UCLA and went to Monsanto in St. Louis, 
where Ward worked as a structural biologist. In 1987 they went to Agouron 
Pharmaceuticals in San Diego. And then in 1995 went cross-country to SmithKline 
Beecham (which became Glaxo SmithKline, merging with GlaxoWellcome).

Ward was very involved with getting IMCA set up as a functional facility for 
pharmaceuticals at Argonne as SmithKline's representative. This experience gave 
him significant credibility in synchrotron macromolecular crystallography, and 
in 2003 he joined the GM/CA-CAT beamlines at the APS to help Bob Fischetti and 
others construct that excellent facility. During this time Cheryl worked at 
Shamrock Structures.

Ward moved to the NIH headquarters in 2007. There he took some responsibility 
for the Protein Structure Initiative, also playing an important role in 
supporting NIH synchrotron facilities. In 2010 he became the branch chief for 
the Structural Genomics and Proteomics Technology Branch in the Division of 
Cell Biology and Biophysics.  He remained in that position through 2017. At the 
2018 NIGMS re-organization Ward went to the Biophysics, Biomedical Technology, 
and Computational Biosciences division as the branch chief for the Biomedical 
Technology Branch.

Ward helped oversee the big NIH-funded, $45 M construction of three major 
beamlines at NSLS-II, a project called ABBIX that ran 2011-2017. In 2017 he 
became program director for NIH support of structural biology beamlines at 
NSLS-II and other DOE synchrotrons.

Many knew Ward for his always calm, reasoned demeanor; he was unflappable, 
resilient, and friendly. He was well read and devoted to his family.


   Robert M. Sweet   E-Dress:  sw...@bnl.gov
   Scientific Advisor, CBMS: The Center for BioMolecular
 Structure at NSLS-II
   Photon Sciences and Biology Dept
   Brookhaven Nat'l Lab.
   Upton, NY  11973 U.S.A.
   Phones: 631 344 3401  (Office)
 631 338 7302  (Mobile)



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

[ccp4bb] [refmac developers] unrestrained anisotropic

[Bernhard Rupp]

Hi Fellows,

 

Q for the Refmac cognosci:

 

Just curious: If I run unrestrained anisotropic refi on 1 A data, some
barely supported ligands display insane and physically impossible ADPs,

worthy of the ORTEP of the century. As cigars penetrate discs, it seems that
Hirschfeld criteria on

aniso ADPs are violated. 

 

Are they also turned off when I select unrestrained - they should remain in
effect regardless of

how bad or unsupported my atoms are? Do I need to change/adjust/optimize the
RBON/RIGU keywrd?

 

Best, BR

 

--

Bernhard Rupp

  http://www.hofkristallamt.org/

  b...@hofkristallamt.org

+1 925 209 7429

+43 676 571 0536

--

Many plausible ideas vanish 

at the presence of thought

--

 




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] Protocol for bacmid transformation in E.coli

[Archit Garg]

Hi Digant,
As Artem mentioned, it's a fairly simple process and it's similar to
plasmid transformation in E. coli. We work extensively with insect cells in
our lab, though we have never used electroporation, the normal chemical
transformation works quite well. Although bacmid requires careful handling
and you will need to isolate it using ethanol/isopropanol precipitation
owing to its size (no usage of columns).

Also, for transformation of your plasmid, you require E. coli cells which
contain bacmid transposition machinery (e.g. DH10Bac cells). More
information on Bac-to-Bac system you can find in this guide from invitrogen-
http://tools.thermofisher.com/content/sfs/manuals/bactobac_man.pdf

Best,
Archit

On Sun, Jul 19, 2020 at 3:12 AM Digant Nayak  wrote:

> Dear all,
>
> Sorry for the off topic question, but i assure you that the long term
> output of the project has structural biology application. I want to
> transform a bacmid into E.coli cell and I was intrigued to find that there
> is no protocol for this available on the internet (i would really
> appreciate it if somebody proves me wrong and directs me to a link). Is it
> similar to transforming a plasmid in E.coli? I would really like to hear
> your experiences on this topic.
>
> Thanks,
> Digant
>
>
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] Sad News

[Fischmann, Thierry]

It’s with much sadness that I learn of Ward’s passing.

To add to Bob’s message: Ward was elected as Chair of the IMCA’s supervisory 
Board. He lead us through some difficult decisions, and always with a great 
sense of humor

Thierry 

> On Jul 18, 2020, at 7:36 AM, Sweet, Robert 
> <27e0eb9d20ec-dmarc-requ...@jiscmail.ac.uk> wrote:
> 
> EXTERNAL EMAIL – Use caution with any links or file attachments.
> 
> I'm writing to acknowledge the passing of Ward Smith during the weekend of 5 
> July. Ward got his PhD with Martha Ludwig at U. of Michigan, and then came to 
> UCLA in 1977 to join Dave Eisenberg’s group as a postdoc. During the course 
> of things, he met Cheryl Janson, a Paul Boyer postdoc, and they were married 
> in 1980.  During his time at UCLA Ward became expert in operation of the 
> Xuong/Hamlin/Nielsen multi-wire system at UCSD and tutored the UCLA users in 
> its use. In 1985 Ward and Cheryl left UCLA and went to Monsanto in St. Louis, 
> where Ward worked as a structural biologist. In 1987 they went to Agouron 
> Pharmaceuticals in San Diego. And then in 1995 went cross-country to 
> SmithKline Beecham (which became Glaxo SmithKline, merging with 
> GlaxoWellcome). 
> 
> Ward was very involved with getting IMCA set up as a functional facility for 
> pharmaceuticals at Argonne as SmithKline's representative. This experience 
> gave him significant credibility in synchrotron macromolecular 
> crystallography, and in 2003 he joined the GM/CA-CAT beamlines at the APS to 
> help Bob Fischetti and others construct that excellent facility. During this 
> time Cheryl worked at Shamrock Structures.
> 
> Ward moved to the NIH headquarters in 2007. There he took some responsibility 
> for the Protein Structure Initiative, also playing an important role in 
> supporting NIH synchrotron facilities. In 2010 he became the branch chief for 
> the Structural Genomics and Proteomics Technology Branch in the Division of 
> Cell Biology and Biophysics.  He remained in that position through 2017. At 
> the 2018 NIGMS re-organization Ward went to the Biophysics, Biomedical 
> Technology, and Computational Biosciences division as the branch chief for 
> the Biomedical Technology Branch. 
> 
> Ward helped oversee the big NIH-funded, $45 M construction of three major 
> beamlines at NSLS-II, a project called ABBIX that ran 2011-2017. In 2017 he 
> became program director for NIH support of structural biology beamlines at 
> NSLS-II and other DOE synchrotrons. 
> 
> Many knew Ward for his always calm, reasoned demeanor; he was unflappable, 
> resilient, and friendly. He was well read and devoted to his family.  
> 
> 
>  Robert M. Sweet   E-Dress:  sw...@bnl.gov
>  Scientific Advisor, CBMS: The Center for BioMolecular
>Structure at NSLS-II
>  Photon Sciences and Biology Dept
>  Brookhaven Nat'l Lab.
>  Upton, NY  11973 U.S.A.
>  Phones: 631 344 3401  (Office)
>631 338 7302  (Mobile)
> 
> 
> 
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
> 
> This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing 
> list hosted by www.jiscmail.ac.uk, terms & conditions are available at 
> https://www.jiscmail.ac.uk/policyandsecurity/
> 
Notice:  This e-mail message, together with any attachments, contains
information of Merck & Co., Inc. (2000 Galloping Hill Road, Kenilworth,
New Jersey, USA 07033), and/or its affiliates Direct contact information
for affiliates is available at 
http://www.merck.com/contact/contacts.html) that may be confidential,
proprietary copyrighted and/or legally privileged. It is intended solely
for the use of the individual or entity named on this message. If you are
not the intended recipient, and have received this message in error,
please notify us immediately by reply e-mail and then delete it from 
your system.




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] protein oligomer

[Nikolay Dobrev]

It really depends from the nature of the protein and if is 
oligomerizing/agregating/forming polymers if any of this is reversible.
On the other side if you are working with one of the fibril forming 
protein it will require optimizaiton on its on as they will form 
naturally polymers.


Do you observed different specises when you analyze your protein by SEC 
or if you are able to perfom DLS?
Additional information regarding your protein will be really helpful for 
more detalied suggestions how to overcome your protein.


Best,
Nikolay

Nikolay Dobrev
Scientific Officer, Protein Expression and Purification Core Facility
EMBL Heidelberg, Meyerhofstraße 1, 69117 Heidelberg, Germany
T +49 6221 387 8633 | M +49 173 684 0532
twitter.com/EMBLorg | facebook.com/embl.org | youtube.com/user/emblmedia
Visit www.embl.org/events for a complete list of all EMBL events.


On 19/07/2020 14:15, S. Mohanty wrote:
Keep the protein concentration low during purification steps along 
with using other anti-aggregation agent/s. Make sure that the pH at 
which you are purifying is not close to the pI of the protein. Until 
completely purified, all purification steps should be performed in a 
cold room if it is a soluble protein.


Smita


On Sunday, July 19, 2020, 04:28:45 AM CDT, Sorin Draga 
 wrote:



I am not sure what you mean by polymer formation. Presuming that you 
have optimized your protein concentration, pH and salt concentration, 
you could try arginine as an anti-aggregation agent in your 
purification (I presume you do FPLC). Have a look at chaotropic agents 
used in protein purification, The answer is generally dependent on the 
protein/proteins you are trying to purify and is not 
necessarily straightforward.


Kinds regards

On Sun, Jul 19, 2020 at 12:08 PM 张士军 <21620150150...@stu.xmu.edu.cn 
> wrote:


Dear all:

Any ideas to decrease protein polymer formation? my protein was
easy to form oligomers and precipitation when do purification,I
have tried add glycerol and DTT,both didn't work. Does anyone has
experience to avoid it happening. Thanks!

Best Regards




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1


--
Nikolay Dobrev
Scientific Officer, Protein Expression and Purification Core Facility
EMBL Heidelberg, Meyerhofstraße 1, 69117 Heidelberg, Germany
T +49 6221 387 8633 | M +49 173 684 0532
twitter.com/EMBLorg | facebook.com/embl.org | youtube.com/user/emblmedia
Visit www.embl.org/events for a complete list of all EMBL events.




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] protein oligomer

[S. Mohanty]

Keep the protein concentration low during purification steps along with using 
other anti-aggregation agent/s. Make sure that the pH at which you are 
purifying is not close to the pI of the protein. Until completely purified, all 
purification steps should be performed in a cold room if it is a soluble 
protein.
Smita  

On Sunday, July 19, 2020, 04:28:45 AM CDT, Sorin Draga 
 wrote:  
 
 I am not sure what you mean by polymer formation. Presuming that you have 
optimized your protein concentration, pH and salt concentration, you could try 
arginine as an anti-aggregation agent in your purification (I presume you do 
FPLC). Have a look at chaotropic agents used in protein purification, The 
answer is generally dependent on the protein/proteins you are trying to purify 
and is not necessarily straightforward.
Kinds regards
On Sun, Jul 19, 2020 at 12:08 PM 张士军 <21620150150...@stu.xmu.edu.cn> wrote:


 Dear all:

 Any ideas to decrease protein polymer formation? my protein was easy to form 
oligomers and precipitation when do purification,I have tried add glycerol and 
DTT,both didn't work. Does anyone has experience to avoid it happening. Thanks!

 Best Regards


To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
  



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] protein oligomer

[Sorin Draga]

I am not sure what you mean by polymer formation. Presuming that you have
optimized your protein concentration, pH and salt concentration, you could
try arginine as an anti-aggregation agent in your purification (I presume
you do FPLC). Have a look at chaotropic agents used in protein
purification, The answer is generally dependent on the protein/proteins you
are trying to purify and is not necessarily straightforward.

Kinds regards

On Sun, Jul 19, 2020 at 12:08 PM 张士军 <21620150150...@stu.xmu.edu.cn> wrote:

> Dear all:
>
> Any ideas to decrease protein polymer formation? my protein was easy to
> form oligomers and precipitation when do purification,I have tried add
> glycerol and DTT,both didn't work. Does anyone has experience to avoid it
> happening. Thanks!
>
> Best Regards
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

[ccp4bb] protein oligomer

[张士军]

Dear all:

Any ideas to decrease protein polymer formation? my protein was easy to form 
oligomers and precipitation when do purification,I have tried add glycerol and 
DTT,both didn't work. Does anyone has experience to avoid it happening. Thanks!

Best Regards



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/