Re: [ccp4bb] Unusual monomer-monomer interface in crystal

[Ivan Shabalin]

Chris,

Let me add my five cents to the discussion.

Processing your data to keep I+ and I- merged separately is a very good 
practice, as noted by Eleanor and Mitch.


Also, I highly recommend using the CMM server 
(https://csgid.org/csgid/metal_sites/) to validate the correctness of 
the metal.


Below are a few publications from our lab that cover these topics:

Characterizing metal-binding sites in proteins with X-ray crystallography.
Nat Protoc 13: 1062-1090. [DOI:10.1038/nprot.2018.018] [Pub Med ID: 
29674755]


CheckMyMetal: a macromolecular metal-binding validation tool.
Acta Crystallogr D Struct Biol 73: 223-233. 
[DOI:10.1107/S2059798317001061] [Pub Med ID: 28291757]


Validation of metal-binding sites in macromolecular structures with the 
CheckMyMetal web server.

Nat Protoc 9: 156-70. [DOI:10.1038/nprot.2013.172] [Pub Med ID: 24356774]

Best,

Ivan


With best regards,
Ivan Shabalin, Ph.D.
Research Scientist,
Department of Molecular Physiology and Biological Physics,
University of Virginia,
1340 Jefferson Park Avenue, Pinn Hall,Room 4223,
Charlottesville, VA 22908
https://www.linkedin.com/in/shabalinig/
https://minorlab.org/person/ivan_s/

On 1/21/20 12:55, Chris Fage wrote:

Dear CCP4BB Users,

I've recently solved the ~2.2 angstrom structure of a protein. In my 
electron density there are unusual monomer-monomer interfaces involving 
pairs of His and Cys residues (see https://ibb.co/wdWBcdk). Note the 
positive Fo-Fc density between the four side chains. As there is not 
adequate space for a water molecule or metal ion, perhaps the Cys 
residues are partially tied up disulfide bonds? However, the 
protein looks to be fully monomeric based on LC-MS measurements. Has 
anyone else observed crystal-driven formation of disulfide bridges?


Aside from this region, there is no extensive interface between 
momoners, and PDBePISA suggests a monomeric state.


Thanks in advance for any advice!

Best wishes,
Chris



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Re: [ccp4bb] Mixed oligomeric states in crystallo

[Diana Tomchick]

Here’s two examples of heterooligomers that crystallized in a lattice with an 
extra monomer of one of the proteins. In both cases this was an unexpected 
result, but easily explained due to the low micro molar affinities for the 
complex.

4PKY
2CJS

Diana

**
Diana R. Tomchick
Professor
Departments of Biophysics and Biochemistry
UT Southwestern Medical Center
5323 Harry Hines Blvd.
Rm. ND10.214A
Dallas, TX 75390-8816
diana.tomch...@utsouthwestern.edu
(214) 645-6383 (phone)
(214) 645-6353 (fax)

On Jan 31, 2020, at 9:23 AM, Kluenemann, Thomas 
mailto:thomas.kluenem...@helmholtz-hzi.de>> 
wrote:


EXTERNAL MAIL

Dear all,

We recently solved a the structure of a small c-type cytochrome. We observed, 
that of the eleven chains in the asymmetric unit ten form 3D domain swapped 
dimers by exchanging an α-helix. The eleventh  chain is present as a monomer. 
Based on the anomalous iron signal and the chain tracing we are sure that no 
chain was missed.
I tried to find other examples in the PDB, were one crystal is made of 
different homo- or heterooligomers.  I only found proteins with partial 
occupied peptide binding sites, which is not what I am looking for. Does anyone 
know of a case were the presence of different homo- or heterooligomers is 
required to form the crystal?

Best regards,
Thomas Klünemann





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Braunschweig | www.helmholtz-hzi.de

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Wissenschaft und Kultur
Geschäftsführung: Prof. Dr. Dirk Heinz; Silke Tannapfel
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[ccp4bb] Fwd: Open scientist position at the EMBL-Grenoble - Project leader for ID30A/MASSIF-1 beamline upgrade

[Andrew MC CARTHY]

 

Dear all, 

Just a quick reminder that the closing date for applications is in one
week (Feb 7th). 

We have an open scientist position at the EMBL-Grenoble for a Project
leader to oversee our contribution to the upgrade of the fully automated
ID30A/MASSIF-1 macromolecular crystallography beamline at the ESRF. This
is an exciting opportunity for an ambitous individual to join a team of
scientists and engineers from the EMBL and ESRF in developing fully
automated data collection protocols to maximise the scientidic potential
of the ESRF Extremely Brilliant Source upgrade. 

Closing date: Feb. 7th 2020. 

Interview date: March 6th, 2020 

You can read more about the position and submit your application at: 

https://www.embl.de/jobs/searchjobs/index.php?ref=GR00139 

Best regards, 

Andrew



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Re: [ccp4bb] Mixed oligomeric states in crystallo

[orly avraham]

Hi,

To my knowledge, the structure of the rhizavidin-biotin complex contains a
hexamer and a monomer, where the monomer forms a dimer using symmetry
operations.
PDB id: 3ew2.

Best,
Orly


On Fri, Jan 31, 2020, 17:33 Kluenemann, Thomas <
thomas.kluenem...@helmholtz-hzi.de> wrote:

> Dear all,
>
>
>
> We recently solved a the structure of a small c-type cytochrome. We
> observed, that of the eleven chains in the asymmetric unit ten form 3D
> domain swapped dimers by exchanging an α-helix. The eleventh  chain is
> present as a monomer. Based on the anomalous iron signal and the chain
> tracing we are sure that no chain was missed.
>
> I tried to find other examples in the PDB, were one crystal is made of
> different homo- or heterooligomers.  I only found proteins with partial
> occupied peptide binding sites, which is not what I am looking for. Does
> anyone know of a case were the presence of different homo- or
> heterooligomers is required to form the crystal?
>
>
>
> Best regards,
>
> Thomas Klünemann
>
>
>
>
>
> --
>
> Helmholtz-Zentrum für Infektionsforschung GmbH | Inhoffenstraße 7 | 38124
> Braunschweig | www.helmholtz-hzi.de
>
> Vorsitzende des Aufsichtsrates: Frau MinDir'in Prof. Dr. Veronika von
> Messling
> Stellvertreter: MinDirig Rüdiger Eichel, Niedersächsisches Ministerium für
> Wissenschaft und Kultur
> Geschäftsführung: Prof. Dr. Dirk Heinz; Silke Tannapfel
> Gesellschaft mit beschränkter Haftung (GmbH)
> Sitz der Gesellschaft: Braunschweig
> Handelsregister: Amtsgericht Braunschweig, HRB 477
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1
>



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[ccp4bb] AW: Mixed oligomeric states in crystallo

[Schreuder, Herman /DE]

Dear Thomas,

The asymmetric unit of the bovine pTAFI structure (3dgv) contains one regular 
dimer and one somewhat disordered monomer. However there is no domain-swapping 
of alpha-helices or other special things.

Best regards,
Herman


Von: CCP4 bulletin board  Im Auftrag von Kluenemann, 
Thomas
Gesendet: Freitag, 31. Januar 2020 16:23
An: CCP4BB@JISCMAIL.AC.UK
Betreff: [EXTERNAL] [ccp4bb] Mixed oligomeric states in crystallo


EXTERNAL : Real sender is 
owner-ccp...@jiscmail.ac.uk

Dear all,

We recently solved a the structure of a small c-type cytochrome. We observed, 
that of the eleven chains in the asymmetric unit ten form 3D domain swapped 
dimers by exchanging an α-helix. The eleventh  chain is present as a monomer. 
Based on the anomalous iron signal and the chain tracing we are sure that no 
chain was missed.
I tried to find other examples in the PDB, were one crystal is made of 
different homo- or heterooligomers.  I only found proteins with partial 
occupied peptide binding sites, which is not what I am looking for. Does anyone 
know of a case were the presence of different homo- or heterooligomers is 
required to form the crystal?

Best regards,
Thomas Klünemann





Helmholtz-Zentrum für Infektionsforschung GmbH | Inhoffenstraße 7 | 38124 
Braunschweig | 
www.helmholtz-hzi.de

Vorsitzende des Aufsichtsrates: Frau MinDir'in Prof. Dr. Veronika von Messling
Stellvertreter: MinDirig Rüdiger Eichel, Niedersächsisches Ministerium für 
Wissenschaft und Kultur
Geschäftsführung: Prof. Dr. Dirk Heinz; Silke Tannapfel
Gesellschaft mit beschränkter Haftung (GmbH)
Sitz der Gesellschaft: Braunschweig
Handelsregister: Amtsgericht Braunschweig, HRB 477



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[ccp4bb] postdoc position

[Alexandra Deaconescu]

A postdoc position is available in my lab. Please see below for more 
information. Thank you.

**

*
*

*POSTDOCTORAL POSITION*

*at Brown University, USA*

The Deaconescu Lab at Brown University has one opening for a 
postdoctoral researcher. The laboratory's interests lie primarily in 
stress responses, with particular emphasis on responses to DNA damage 
and the mechanochemistry of DNA-based motors. We utilize a combination 
of biochemical, biophysical and structural techniques. (e.g. X-ray 
crystallography, small-angle X-ray scattering, transmission electron 
microscopy). For examples of work, please see Dorich et al., /Genes Dev/ 
(2019), Vemu et al. /Science /(2018), Le TT et al. /Cell/ (2018), 
Deaconescu et al. /Photochemistry and Photobiology /(2017), Kutter et 
al. /JMB/ (2016), Szyk et al, /Cell/ (2014), Deaconescu et al., /PNAS/ 
(2012), Szyk et al., /NSMB/ (2012) and Deaconescu et al., /TIBS/ (2013).


A successful candidate should have an established track-record of 
publications in peer-reviewed journals; solid experience with protein 
biochemistry (including their purification from E.coli/yeast expression, 
functional characterization and assay development).*_Prior knowledge of 
crystallography and/or single-particle electron microscopy is highly 
desirable_*. Must be highly motivated and work independently as well as 
in a team. Excellent spoken and written English are required. New Ph.D. 
graduates are encouraged to apply.


*Interested candidates should send a CV, one page research experience 
summary and contact information for three references to 
**alexandra_deacone...@brown.edu* 
*.*Salary and starting date are 
negotiable.Please write Postdoctoral Candidate in the e-mail subject 
header.Brown University, an Ivy League school, is located less than one 
hour away by train from Boston.Lab webpage: https://deaconesculab.com


--
Alexandra Deaconescu, B.E., Ph.D.
Assistant Professor
Brown University

Office: (401) 863-3215
Wet Lab: (401) 863-6729
Computational Lab: (401) 863-7031

For Mail:
Laboratories of Molecular Medicine
70 Ship St. GE-4
Providence, RI 02903

For Courier:
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Brown University
70 Ship St., Chestnut St. Loading Dock
Providence, RI 02903

Website: www.deaconesculab.com

Admin
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Email: richard_waugh[at]brown.edu
Mailing Address:
Box G-E, Brown University,
Providence, RI 02912-G
Telephone: 401-863-7446

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[ccp4bb] Mixed oligomeric states in crystallo

[Kluenemann, Thomas]

Dear all,

We recently solved a the structure of a small c-type cytochrome. We observed, 
that of the eleven chains in the asymmetric unit ten form 3D domain swapped 
dimers by exchanging an α-helix. The eleventh  chain is present as a monomer. 
Based on the anomalous iron signal and the chain tracing we are sure that no 
chain was missed.
I tried to find other examples in the PDB, were one crystal is made of 
different homo- or heterooligomers.  I only found proteins with partial 
occupied peptide binding sites, which is not what I am looking for. Does anyone 
know of a case were the presence of different homo- or heterooligomers is 
required to form the crystal?

Best regards,
Thomas Klünemann





Helmholtz-Zentrum für Infektionsforschung GmbH | Inhoffenstraße 7 | 38124 
Braunschweig | www.helmholtz-hzi.de

Vorsitzende des Aufsichtsrates: Frau MinDir'in Prof. Dr. Veronika von Messling
Stellvertreter: MinDirig Rüdiger Eichel, Niedersächsisches Ministerium für 
Wissenschaft und Kultur
Geschäftsführung: Prof. Dr. Dirk Heinz; Silke Tannapfel
Gesellschaft mit beschränkter Haftung (GmbH)
Sitz der Gesellschaft: Braunschweig
Handelsregister: Amtsgericht Braunschweig, HRB 477



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[ccp4bb] Electron microscopy support position @ CSSB, Hamburg

[Thomas White]

Hi everyone,

The CryoEM facility at CSSB in Hamburg is growing
(https://www.cssb-hamburg.de/facilities/cryo_em/index_eng.html)

We are currently looking for an EM support scientist to complement our
Team.  A job description is pasted below - however, we are totally
aware, that everyone contributes her/his own expertise, and that this
may change some of the responsibilities described.

For more information, or any other questions, please contact:
Carolin Seuring 


Electron Microscopy Support (m/f/D) - Salary Level 13 TV-L -

The position commences as soon as possible and is permanent. This
position is also suitable for part time employment.

The Centre for Structural Systems Biology (CSSB) was created with the
mission to use cutting edge technologies and methods to investigate how
pathogens infect humans. Part of this mission are the CSSB Core
Facilities that provide access to technology and research services for
bio-sciences. Alongside these facilities, the multi-user facility for
electron cryo-microscopy (cryoEM) has started its operation in 2017,
and is positioned on the Science Campus Bahrenfeld, being also home to
the Deutsches Elektronen-Synchrotron DESY. Our facility is currently
the most advanced centre for cryoEM in the north of Germany housing
five electron microscopes: two Titan Krios instruments equipped with
phase plates, energy filters, K3 and Falcon 3 direct detectors, a Talos
Arctica with a phase plate and a Falcon 3 direct detector, a Talos
L120C and an Aquilos FIB cryo-SEM for preparing lamellae of thick
samples, such as vitrified cells or crystals. Auxiliary equipment and
nearby lab space for sample preparation and cell culture work complete
the workflows. We primarily focus on single particle analysis and
electron cryo-tomography. The facility is set up to handle biological
samples up to German biosafety level 2. 

Nature of the job

85% equipment set-up and operation, equipment maintenance, 15%
user-training & assistance:

Responsible for the day-to-day running and maintenance of the electron
microscopes and associated sample preparation equipment

Manage and operate the cryoEM instruments and associated equipment,
this includes first-fault diagnosis, and maintenance work

Assist in improving set-ups for the different cryoEM modalities (SPA,
Tomo) and the implementation of additional workflows (correlative
microscopy, microED)

Work with our computing scientist to ensure a seamless interface
between data collection and data processing

Establish and implement schedules for microscope hardware and software
maintenance, and support alignments and performance checks, to ensure
that the facility is operating at peak performance

Coordinate repairs with microscope supplier’s service engineers

Work with in house engineers and other facilities provide instructions,
assistance and consultation to faculty, students, staff members
regarding the TEM equipment and optimal processes

Perform other related duties incidental to the work described above

Your Profile

Experience in cryoEM (hardware and/or software), X-ray
technology, imaging detectors, or computational work at the interface
hardware/software

Experience in cryoEM & understanding the basics of
electron optics and vacuum systems

Previous experience in microscope control and acquisition software
packages for single particle analysis or tomography or one of the
following programming interfaces/languages: Python, C++, Java, LabView,
SerialEM

Good English language skills (both written and oral)

Experience with managing, analyzing, and improving complex workflows

Demonstrated record keeping and organization skills

Ability to proactively plan maintenance work and increase microscope
uptime & performance

Attention to detail

A desire to learn, and a willingness to share knowledge

Self-driven and organized

Your responsibilities within the team can be adjusted to best fit your
specific skill-set.

For more information please visit:
https://www.uni-hamburg.de/uhh/stellenangebote/technisches-bibliotheks-und-verwaltungspersonal/technik-und-service-und-sonstiges/17-02-20-tvp-602-1-en.pdf



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[ccp4bb] CryoEM and beyond - an exciting postdoc opportunity in Exeter

[Daum, Bertram]

Dear all,

An exciting and highly interdisciplinary ERC-funded postdoc position is now 
available in our lab at the Living Systems Institute in Exeter, UK.

https://www.exeter.ac.uk/livingsystems/

We are looking for an enthusiastic researcher to investigate the structure and 
function of S-layers by cryoEM, with the aim to implement these into innovative 
microrobotic devices.

https://www.exeter.ac.uk/livingsystems/research/biohybrid-microrobots/

If you are interested, please apply here.

https://jobs.exeter.ac.uk/hrpr_webrecruitment/wrd/run/ETREC107GF.open?VACANCY_ID=021881RlTr&WVID=3817591jNg&LANG=USA

The closing date for applications is 12th March 2020.

Cheers,
Bertram

--
Dr Bertram Daum
Senior Research Fellow
University of Exeter

+441392727455
www.exeter.ac.uk

Living Systems Institute
Stocker Road
Exeter
Devon
EX4 4QD
United Kingdom

https://twitter.com/DaumLaboratory
https://www.exeter.ac.uk/livingsystems/team/profile/index.php?web_id=Bertram_Daum
http://www.bristol.ac.uk/wolfson-bioimaging/equipment/cryo-em/






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