[ccp4bb] Large scale insect cell expression

2018-12-11 Thread Joseph Brock 
Dear ccp4 community,


I'll soon start my own lab and together with other colleagues, would like to 
establish a facility to do large scale insect cell culture for crystallography. 
Has anyone had any experience with the wave bag systems for this purpose (for 
instance from GE)? I have heard that culture infection can be a problem with 
this system.


Any advice or other helpful tips in this regard would be much appreciated.


Best regards,

Joseph.




Joseph S. Brock | PhD
Researcher
Drew Lab
Department of Biochemistry and Biophysics
Stockholm University



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Re: [ccp4bb] suggestions for cryoprotectant

2018-10-20 Thread Joseph Brock 
Dear Friduous,


This is also a good paper to look at once you know your crystals diffract at 
room temperature:

https://www.ncbi.nlm.nih.gov/pubmed/10944336


Many of the salts in your condition will be cryo-protective if you increase 
their concentration to saturated levels.


Cheers,

Joseph.


Joseph S. Brock | PhD
Researcher
Drew Lab
Department of Biochemistry and Biophysics
Stockholm University

From: CCP4 bulletin board  on behalf of CCP4BB automatic 
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Sent: Sunday, 21 October 2018 1:00:06 AM
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Subject: CCP4BB Digest - 19 Oct 2018 to 20 Oct 2018 (#2018-267)

There are 5 messages totaling 760 lines in this issue.

Topics of the day:

  1. suggestions for cryoprotectant (4)
  2. RapiData at SSRL 2019



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--

Date:Fri, 19 Oct 2018 17:23:49 -0700
From:Christine Gee 
Subject: Re: suggestions for cryoprotectant

Dear Fridous

Have a look at https://www.ncbi.nlm.nih.gov/pubmed/14646118
[http://www.ncbi.nlm.nih.gov/coreutils/img/pubmed256blue.png]

Malonate: a versatile cryoprotectant and stabilizing solution for salt-grown 
macromolecular crystals. - PubMed - 
NCBI
www.ncbi.nlm.nih.gov
Acta Crystallogr D Biol Crystallogr. 2003 Dec;59(Pt 12):2356-8. Epub 2003 Nov 
27.


Malonate as a cryoprotectant

Or https://www.ncbi.nlm.nih.gov/pubmed/20606259
[http://www.ncbi.nlm.nih.gov/coreutils/img/pubmed256blue.png]

Cryoprotection properties of salts of organic acids: a case study for a 
tetragonal crystal of HEW lysozyme. - PubMed - 
NCBI
www.ncbi.nlm.nih.gov
Acta Crystallogr D Biol Crystallogr. 2010 Jul;66(Pt 7):789-96. doi: 
10.1107/S0907444910015416. Epub 2010 Jun 19.



Regards
Christine

On Fri, Oct 19, 2018 at 2:57 PM Firdous Tarique 
wrote:

> Dear members
>
> I have got beautiful crystal hits in SaltRx screens which are not
> diffracting to a good resoultion. All of them are salt based condition and
> I am not able to formulate a good cryoprotectant for these crystals. I also
> think that in my case the poor resolution is due to a poor cryoprotectant
> selection.
>
> The conditions are as follows:
>
> 1> 4M Ammonium Acetate 100mM Bis Tris Propane pH 7.0
> 2>0.5M KCN 100mM Tris pH8.5
> 3>1.5M LiSo4 100mM Bris Tris Propane pH 7.0
> 4>4M Sodium Nitrate 100mM Tris pH8.5
> 5>1.5M Sodium Nitrate 100mM Sodium acetate pH 4.6
>
> There are few more conditions but so far not able to see good diffraction
> with using lower peg and glycerol based cryoprotectants.
>
> Can anybody suggest me good cryos conditions for salt based
> crystallization conditions or anything good for SaltRx crystallization hits.
>
> Thanks
>
> Firdous
>
> --
>
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Date:Sat, 20 Oct 2018 02:21:41 +
From:"Smith, Clyde" 
Subject: RapiData at SSRL 2019

Hi everyone,

Applications are now open for the 2019 edition of RapiData at SSRL. RapiData is 
a practical course in macromolecular X-ray diffraction data collection, data 
processing and structure solution. The aim of the RapiData course is to educate 
and train young scientists in data collection and processing methods at 
synchrotron beamlines, using state-of-the-art software and instrumentation. The 
course will be held at the Stanford Synchrotron Radiation Lightsource (SSRL) 
located on the SLAC National Accelerator Laboratory campus in Menlo Park, CA, 
from May 5-10 2019. Course organizers for 2019 are Silvia Russi, Clyde Smith 
and Jeney Wierman.

The course will comprise hands-on experiments at the SSRL beamlines, software 
tutorials, and lectures on the following topics:
Specimen preparation, Data collection strategies, X-ray light sources, X-ray 
detectors, Data reduction, Structure solution by MAD, SAD and Molecular 
Replacement, Complementary methods (spectroscopy and small angle scattering)

Please visit http://smb.slac.stanford.edu/news/rapidata/rapidata-2019/ and use 
the links on the "registration" page to apply for the course. The application 
deadline is December 31 2018 Successful applicants will be notified early in 
January 2019 and invited to register and book accommodation at the SLAC 
Guesthouse at that time. A limited amount of travel support funding may be 
availa

[ccp4bb] Post-doc in membrane protein structural biology, Karolinska Institute.

2017-06-01 Thread Joseph Brock 
Dear colleagues,

Please see the information below regarding an opening in our research group. 
Please share with anyone who might be interested.

Best regards,

-Joseph.



A two-year post-doctoral position is available at the Department of Medical 
Biochemistry and Biophysics (MBB), Karolinska Institutet, Stockholm. The 
project deals primarily with over-expression, purification, crystallisation and 
structure determination of human membrane proteins involved in the generation 
of lipid mediators of inflammation. The successful candidate will join a 
multi-disciplinary research team and the position represents an opportunity for 
a postdoctoral scientist to develop their expertise as well as broaden their 
experience. The major interests of the lab are in membrane protein structure 
and function. We combine structural techniques such as macromolecular 
crystallography with biophysical, biochemical and computational methods to 
investigate the structural basis of catalysis. The lab has all necessary 
equipment and access to facilities needed for the project, including a core 
facility for protein science. We have periodic access to beamlines at ESRF 
(France), Diamond (UK), BESSY and DESY (Germany) and also have access to 
state-of-art cryo-EM facilities.

Qualifications:
The candidate should hold a PhD conferred within the previous two years (or 
expect conferral in the near future), be motivated to independently solve 
problems and a have a strong background in biochemistry or structural biology. 
Experience with protein expression, purification and biochemical 
characterisation is required.  Prior experience in membrane protein 
biochemistry and computational skills such as a familiarity with the LINUX 
operating system will be added advantages.

The application should contain a single PDF document including:
·   Letter of intention stating the scientific interests, technical skills, 
and previous experience of the candidate.
·   CV including date and field of (expected) graduation and list of 
publications, with the contact information of at least two references.

This should be sent to joseph.br...@ki.se and jesper.haeggst...@ki.se.

The deadline for application is the 22nd of June, 2017.



Joseph Brock | PhD
Division of Physiological Chemistry II
Department of Medical Biochemistry and Biophysics
Karolinska Institutet
Scheeles väg 2
SE-171 77 Stockholm, Sweden

[ccp4bb] Post-doc in membrane protein structural biology, Karolinska Institute.

2017-04-05 Thread Joseph Brock 
Dear colleagues,

Please see the information below regarding an opening in our research group. 
Please share with anyone who might be interested.

Thanks!

-Joseph.



A two-year post-doctoral position is available at the Department of Medical 
Biochemistry and Biophysics (MBB), Karolinska Institutet, Stockholm. The 
project deals primarily with over-expression, purification, crystallisation and 
structure determination of human membrane proteins involved in the generation 
of lipid mediators of inflammation. The successful candidate will join a 
multi-disciplinary research team and the position represents an opportunity for 
a postdoctoral scientist to develop their expertise as well as broaden their 
experience. The major interests of the lab are in membrane protein structure 
and function. We combine structural techniques such as macromolecular 
crystallography with biophysical, biochemical and computational methods to 
investigate the structural basis of catalysis. The lab has all necessary 
equipment and access to facilities needed for the project, including a core 
facility for protein science. We have periodic access to beamlines at ESRF 
(France), Diamond (UK), BESSY and DESY (Germany).

Qualifications:
The candidate should hold a PhD conferred within the previous two years (or 
expect conferral in the near future), be motivated to independently solve 
problems and a have a strong background in biochemistry or structural biology. 
Experience with protein expression, purification and biochemical 
characterisation is required.  Prior experience in membrane protein 
biochemistry and computational skills such as a familiarity with the LINUX 
operating system will be added advantages.

The application should contain a single PDF document including:

*   Letter of intention stating the scientific interests, technical skills, 
and previous experience of the candidate.

*   CV including date and field of (expected) graduation and list of 
publications, with the contact information of at least two references.

This should be sent to joseph.br...@ki.se and jesper.haeggst...@ki.se.

The deadline for application is the 28th of April, 2016.




Joseph Brock | PhD
Division of Physiological Chemistry II
Department of Medical Biochemistry and Biophysics
Karolinska Institutet
Scheeles väg 2
SE-171 77 Stockholm, Sweden

Re: [ccp4bb] Post-doctoral position in membrane structural biology, Karolinska Institute.

2017-02-21 Thread Joseph Brock 
Ps - application deadline is the 13th of March, 2017.

Best,

-Joseph.

Joseph Brock | PhD
Division of Physiological Chemistry II
Department of Medical Biochemistry and Biophysics
Karolinska Institutet
Scheeles väg 2
SE-171 77 Stockholm, Sweden


From: Joseph Brock
Sent: Tuesday, 21 February 2017 4:52 PM
To: ccp4bb@jiscmail.ac.uk
Subject: Post-doctoral position in membrane structural biology, Karolinska 
Institute.

Dear colleagues,

Please see the information below regarding an opening in our research group. 
Please share with anyone who might be interested.

Thanks!

-Joseph.



A two-year post-doctoral position is available at the Department of Medical 
Biochemistry and Biophysics (MBB), Karolinska Institutet, Stockholm. The 
project deals primarily with over-expression, purification, crystallisation and 
structure determination of human membrane proteins involved in the generation 
of lipid mediators of inflammation. The successful candidate will join a 
multi-disciplinary research team and the position represents an opportunity for 
a postdoctoral scientist to develop their expertise as well as broaden their 
experience. The major interests of the lab are in membrane protein structure 
and function. We combine structural techniques such as macromolecular 
crystallography with biophysical, biochemical and computational methods to 
investigate the structural basis of catalysis. The lab has all necessary 
equipment and access to facilities needed for the project, including a core 
facility for protein science. We have periodic access to beamlines at ESRF 
(France), Diamond (UK), BESSY and DESY (Germany).

Qualifications:
The candidates should hold a PhD, be motivated to independently solve problems 
and a have a strong background in biochemistry or structural biology. 
Experience with protein expression, purification and biochemical 
characterisation is required.  Prior experience in membrane protein 
biochemistry and computational skills such as a familiarity with the LINUX 
operating system will be added advantages.

The application should contain a single PDF document including:

*   Letter of intention stating the scientific interests, technical skills, 
and previous experience of the candidate.

*   CV including date and field of (expected) graduation and list of 
publications, with the contact information of at least two references.

This should be sent to joseph.br...@ki.se and jesper.haeggst...@ki.se

Joseph Brock | PhD
Division of Physiological Chemistry II
Department of Medical Biochemistry and Biophysics
Karolinska Institutet
Scheeles väg 2
SE-171 77 Stockholm, Sweden

[ccp4bb] Post-doctoral position in membrane structural biology, Karolinska Institute.

2017-02-21 Thread Joseph Brock 
Dear colleagues,

Please see the information below regarding an opening in our research group. 
Please share with anyone who might be interested.

Thanks!

-Joseph.



A two-year post-doctoral position is available at the Department of Medical 
Biochemistry and Biophysics (MBB), Karolinska Institutet, Stockholm. The 
project deals primarily with over-expression, purification, crystallisation and 
structure determination of human membrane proteins involved in the generation 
of lipid mediators of inflammation. The successful candidate will join a 
multi-disciplinary research team and the position represents an opportunity for 
a postdoctoral scientist to develop their expertise as well as broaden their 
experience. The major interests of the lab are in membrane protein structure 
and function. We combine structural techniques such as macromolecular 
crystallography with biophysical, biochemical and computational methods to 
investigate the structural basis of catalysis. The lab has all necessary 
equipment and access to facilities needed for the project, including a core 
facility for protein science. We have periodic access to beamlines at ESRF 
(France), Diamond (UK), BESSY and DESY (Germany).

Qualifications:
The candidates should hold a PhD, be motivated to independently solve problems 
and a have a strong background in biochemistry or structural biology. 
Experience with protein expression, purification and biochemical 
characterisation is required.  Prior experience in membrane protein 
biochemistry and computational skills such as a familiarity with the LINUX 
operating system will be added advantages.

The application should contain a single PDF document including:

*   Letter of intention stating the scientific interests, technical skills, 
and previous experience of the candidate.

*   CV including date and field of (expected) graduation and list of 
publications, with the contact information of at least two references.

This should be sent to joseph.br...@ki.se and jesper.haeggst...@ki.se

Joseph Brock | PhD
Division of Physiological Chemistry II
Department of Medical Biochemistry and Biophysics
Karolinska Institutet
Scheeles väg 2
SE-171 77 Stockholm, Sweden

Re: [ccp4bb] POINTLESS error: "NormaliseInRuns::apply Unused run 0"

2017-01-23 Thread Joseph Brock 
Hi Elanor and bb,

I realised this was due to me creating a .HKL file with XDS from Eiger data 
with a DATA_RANGE that was not a multiple of 50!

Apologies for any trouble!

All the best,

Joseph.


Joseph Brock | PhD
Division of Physiological Chemistry II
Department of Medical Biochemistry and Biophysics
Karolinska Institutet
Scheeles väg 2
SE-171 77 Stockholm, Sweden


From: Eleanor Dodson [eleanor.dod...@york.ac.uk]
Sent: Saturday, 21 January 2017 1:07 PM
To: Joseph Brock
Cc: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] POINTLESS error: "NormaliseInRuns::apply Unused run 0"

can you send the whole log file?
Somehow the XDS input isnt agreeing with pointless expectations, so I suspect 
no reflections have been read
Eleanor

On 20 January 2017 at 23:41, Joseph Brock 
mailto:joseph.br...@ki.se>> wrote:
Dear ccp4 experts,

I am trying to use the ccp4i Pointless, Aimless, Ctruncate application to 
convert a XDS.HKL file. This usually works fine, however I suddenly have the 
task immediately fail with the error message below pasted from the bottom of 
the logfile.

If anyone can translate this error message for me I would be very grateful.

I am running up to date ccp4 (7.0.027) on Ubuntu 14.04.



***
* Information from CCP4Interface script
***
The program run with command: /usr/local/bin/ccp4-7.0/bin/pointless HKLOUT 
"/tmp/joe/jsb_32_2_mtz.tmp"
has failed with error message
NormaliseInRuns::apply Unused run 0
***


#CCP4I TERMINATION STATUS 0 "NormaliseInRuns::apply Unused run 0"
#CCP4I TERMINATION TIME 21 Jan 2017  00:18:31
#CCP4I MESSAGE Task failed


Thanks in advance for the help.

Cheers,
Joe.

Joseph Brock | PhD
Division of Physiological Chemistry II
Department of Medical Biochemistry and Biophysics
Karolinska Institutet
Scheeles väg 2
SE-171 77 Stockholm, Sweden

[ccp4bb] POINTLESS error: "NormaliseInRuns::apply Unused run 0"

2017-01-20 Thread Joseph Brock 
Dear ccp4 experts,

I am trying to use the ccp4i Pointless, Aimless, Ctruncate application to 
convert a XDS.HKL file. This usually works fine, however I suddenly have the 
task immediately fail with the error message below pasted from the bottom of 
the logfile.

If anyone can translate this error message for me I would be very grateful.

I am running up to date ccp4 (7.0.027) on Ubuntu 14.04.



***
* Information from CCP4Interface script
***
The program run with command: /usr/local/bin/ccp4-7.0/bin/pointless HKLOUT 
"/tmp/joe/jsb_32_2_mtz.tmp"
has failed with error message
NormaliseInRuns::apply Unused run 0
***


#CCP4I TERMINATION STATUS 0 "NormaliseInRuns::apply Unused run 0"
#CCP4I TERMINATION TIME 21 Jan 2017  00:18:31
#CCP4I MESSAGE Task failed


Thanks in advance for the help.

Cheers,
Joe.

Joseph Brock | PhD
Division of Physiological Chemistry II
Department of Medical Biochemistry and Biophysics
Karolinska Institutet
Scheeles väg 2
SE-171 77 Stockholm, Sweden

Re: [ccp4bb] structure based superposition

2011-08-18 Thread Joseph Brock 

Hi Suda,

You can do this in Coot by loading the two structures, then using the tool 
Calculate-> SSM superpose.

The rmsd value and sequence identity should be reported within your terminal 
window.

Hope that helps.

-Joe

Date: Thu, 18 Aug 2011 14:40:32 +0530
From: biobud...@gmail.com
Subject: [ccp4bb] structure based superposition
To: CCP4BB@JISCMAIL.AC.UK

Dear all,

I would like to know the tools/servers/programs that can be used for structure 
based superposition of two or more proteins. Please help me out..!!

Thanks,

-Suda

[ccp4bb] Seeding and protease questions

2009-09-15 Thread Joseph Brock 

Hi everyone,

I have two conditions that I want to try and improve by microseeding. I have 
been reading the literature on the subject and am a bit confused as to what 
exactly is the "stabilizing mother liquor" that is used in the serial dilution 
of the seed stock. Is this just purified protein in its stabilizing buffer? Or 
a mixture of this and crystallization solution? I would also like to know what 
the general opinion is on WHEN to add these seed stocks to crystallization 
drop. Is addition generally more effective upon setup or after the drops have 
had a chance to equilibrate?

Also, one of these conditions takes a LONG time to produce crystals (about 3 
months). I understand this may be due to the slow digestion of my protein by 
low levels of co-purified proteases, whose slow nibbling eventually produce a 
truncated form that is more conducive to forming crystals. Thus, the process 
can be accelerated by the addition of something like pepsin to the 
crystallization drop. I was hoping that someone could offer advice as to what 
the optimal protease/concentration to use is such experiments?

Many thanks in advance for the advice and everything I have learnt from this 
community in the past.

Best regards,

Joe
PhD Student
Research School of Chemistry
Australian National University

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