Re: [ccp4bb] Nitril groups not present on compounds

[Holton, James M]

10% of which beam?

-James Holton
MAD Scientist

On Feb 26, 2019, at 1:38 PM, Maria Håkansson 
mailto:maria.hakans...@saromics.com>> wrote:

Dear all,

We have experienced nitril groups not present in the electron density
after data collection on compounds bound to different proteins.

In one of the projects the compound was tested by MS to check that
the nitril is present in solution before data collection (crystals used to for 
MS analysis).
The nitril should be present since the molecular weight of the compound is the 
same.

Also the crystals have been exposed using 10% of the beam (instead of 100%) to 
try to make
the effect of radiation damage as small as possible. Still the nitril is not 
present.

In both cases the nitril atoms have been modeled with high temperature factors 
but
the electron density is missing, see parts of the compound in the image below 
contoured at 1 sigma level and in a 1.1 Å electron density map.

Has anyone else experienced this?

Best regards,
Maria



Maria Håkansson, PhD, Crystallization Facility Manager
Principal Scientist

SARomics Biostructures AB
Medicon Village
SE-223 81 Lund, Sweden

Mobile: +46 (0)76 8585706
Web: www.saromics.com







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Re: [ccp4bb] Nitril groups not present on compounds

[Mark J van Raaij]

Hi Maria,

from the picture, it looks like disorder to me. Note that the three non-H atoms 
at the top of the ring also have significantly less density than the five non-H 
atoms that are closer to the protein. Perhaps the compounds is just quite 
wobbly in it's binding site.

Mark J van Raaij
Dpto de Estructura de Macromoleculas
Centro Nacional de Biotecnologia - CSIC
calle Darwin 3
E-28049 Madrid, Spain
tel. (+34) 91 585 4616
http://wwwuser.cnb.csic.es/~mjvanraaij
Section Editor of Acta Crystallographica F, Structural Biology Communications
http://journals.iucr.org/f/


> On 26 Feb 2019, at 13:38, Maria Håkansson  
> wrote:
> 
> Dear all,
> 
> We have experienced nitril groups not present in the electron density
> after data collection on compounds bound to different proteins.
> 
> In one of the projects the compound was tested by MS to check that
> the nitril is present in solution before data collection (crystals used to 
> for MS analysis).
> The nitril should be present since the molecular weight of the compound is 
> the same.
> 
> Also the crystals have been exposed using 10% of the beam (instead of 100%) 
> to try to make 
> the effect of radiation damage as small as possible. Still the nitril is not 
> present.
> 
> In both cases the nitril atoms have been modeled with high temperature 
> factors but
> the electron density is missing, see parts of the compound in the image below 
> contoured at 1 sigma level and in a 1.1 Å electron density map.
> 
> Has anyone else experienced this?
> 
> Best regards,
> Maria
>  
> 
> 
> Maria Håkansson, PhD, Crystallization  13.34.17.png>Facility Manager
> Principal Scientist
> 
> SARomics Biostructures AB
> Medicon Village
> SE-223 81 Lund, Sweden
> 
> Mobile: +46 (0)76 8585706
> Web: www.saromics.com 
> 
> 
> 
> 
> 
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1 
> 



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[ccp4bb] Postdoctoral position in structural biology of influenza virus transcription Cusack group, EMBL Grenoble

[Stephen Cusack]

Dear All,

    We are looking for a highly motivated and ambitious individual to 
progress our understanding of the mechanism of transcription by 
influenza virus RNA-dependent RNA polymerase. The work is in the frame 
of the ANR-funded FluTranscript project that brings together the 
complementary expertise of the groups of Stephen Cusack (EMBL Grenoble, 
project coordinator) and Nadia Naffakh (Institut Pasteur) in structural 
biology and molecular virology, respectively. The project follows on 
from a recent joint publication (Structural basis of an essential 
interaction between influenza polymerase and Pol II CTD. Lukarska et al, 
Nature 2017) and aims to reconstitute and determine the structure of 
functional transcription complexes. A secondary goal is to use the 
acquired structural information to develop new approaches to 
anti-influenza drug development.


    The Cusack group uses X-ray crystallography and cryo 
electron-microscopy to study the structural biology of protein-RNA 
complexes involved in RNA virus replication, innate immunity and 
cellular RNA metabolism. Group website: 
https://www.embl.fr/research/unit/cusack/index.html


    The successful candidate should have a PhD in Molecular Biology or 
related field and a strong background and interest in biochemistry and 
structural biology.
We are looking for a creative and ambitious person with good 
communication skills and keen to work on a challenging project.


    Experience in insect cell expression systems, reconstitution of 
protein-RNA complexes, X-ray crystallography or cryo-electron microscopy 
would be particularly beneficial.


    EMBL is one of the world’s leading research organizations in the 
life sciences. The Headquarters Laboratory is located in Heidelberg, 
with additional sites in Barcelona, Grenoble, Hamburg, Hinxton, and Rome.


    EMBL Grenoble has excellent access to state-of-the-art structural 
biology technologies with the in house EM facility being equipped with a 
T12 microscope and 200kV Glacios/Falcon III . Additionally, we have 
access to the Titan Krios at the adjacent ESRF as well as those in EMBL 
Heidelberg. Other facilities on the Grenoble Campus include the ESRF 
synchrotron X-ray beamlines, neutron scattering at the ILL, high-field 
NMR at the IBS as well as high-throughput crystallization platform, 
mammalian and insect cell facilities, biophysical platform and confocal 
microscopy.


You can find out more about this vacancy at:
https://www.embl.fr/jobs/searchjobs/index.php?ref=GR00133

or by contacting Stephen Cusack: cus...@embl.fr

Please apply online through www.embl.org/

Stephen Cusack

--

**
Dr. Stephen Cusack, FRS
Head of Grenoble Outstation of the European Molecular Biology Laboratory (EMBL)
Group leader in structural biology of protein-RNA complexes and viral proteins
**

Email:  cus...@embl.fr  
Website: http://www.embl.fr 
Tel:(33) 4 76 20 7238Secretary (33) 4 76 20 7123

Fax:(33) 4 76 20 7199   
Postal address:   EMBL Grenoble Outstation, 71 Avenue des Martyrs, CS 90181, 
38042 Grenoble Cedex 9, France
Delivery address: EMBL Grenoble Outstation, Polygone Scientifique, 71 Avenue 
des Martyrs, 38000 Grenoble, France
**



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[ccp4bb] CSMB Postdoctoral Research Associate, Oak Ridge National Laboratory

[O'Neill, Hugh]

Postdoctoral Research Associate at the Center for Structural Molecular Biology, 
Oak Ridge National Laboratory

The Center for Structural Molecular Biology (CSMB) has a postdoctoral fellow 
position available to work in the area of structural biology/experimental 
biophysics. In this position, you will be expected to contribute to the 
development of methods for investigating the structure of biomacromolecular 
complexes and assemblies using neutron scattering in combination with other 
experimental approaches. Desirable skills include expertise in biological 
structure determination using scattering and/or diffraction techniques, protein 
expression and purification, and molecular simulations. Prior experience in 
characterizing membrane protein complexes is preferred. You will be involved in 
structural measurements on a range of macromolecular targets and data analysis 
using state-of-the-art techniques and facilities available at the High Flux 
Isotope Reactor (HFIR), the United States' highest flux reactor-based neutron 
source, and the Spallation Neutron Source (SNS), the world's most intense 
pulsed accelerator-based neutron source (http://neutrons.ornl.gov). You will 
also have access to a suite of biophysical characterization tools, as well as 
external synchrotron facilities. The CSMB is a premier research center focused 
on the characterization of biological systems with relevance to environmental, 
biotechnological and biomedical systems. It supports the biological small-angle 
neutron scattering (Bio-SANS) instrument at HFIR and the bio-deuteration 
laboratory, dedicated to the production of labeled biomolecules, at the SNS. It 
has an outstanding group of scientists and technical staff focused on basic and 
applied research, and strong ties to the Oak Ridge Leadership Computing 
Facility and Center for Nanophase Materials Science.


For informal enquiries please contact Hugh O'Neill 
(oneil...@ornl.gov)
For further details and to apply, please see link below:
https://jobs.ornl.gov/job/Oak-Ridge-Postdoctoral-Research-Associate-Structural-Molecular-Biology-TN-37831/539682100/




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[ccp4bb] Reminder: Call for MX beamtime proposals at HZB, BESSY II, deadline March 01, 2019

[Manfred S. Weiss]

Dear all,

I would just like to remind you of the upcoming deadline.

Best wishes
Manfred, on behalf of the HZB-MX team




the next MX-proposal application deadline: March 01, 2019 is approaching
https://www.helmholtz-berlin.de/user/beamtime/proposals/index_en.html

As usual, all proposals will be handled with the new version of GATE,
GATE2.0, https://www.helmholtz-berlin.de/pubbin/hzbgate

Hereby we would like to invite the submission of new proposals for
MX-beamtime at the HZB-MX beamlines for the next beam time period
(09/2019-02/2020).

In order to apply for beamtime, please register at the HZB on-line
access tool "GATE" (https://www.helmholtz-berlin.de/pubbin/hzbgate)
and submit a new beam time application proposal.

With the new GATE version, the proposal categories have been improved.
We now expect from each research group only ONE proposal, which can
contain up to 20 individual projects.

HZB provides MX-beamtime at the three MX-beamlines BL14.1, BL14.2
and BL14.3. The three beamlines are equipped with state-of-the-art
instrumentation and are currently the most productive MX-stations in
Germany with over 300 PDB depositions annually. Beamtime is granted
based on the reviewed proposals and on reports from previous research
activities. Please make sure to include them if available.

Experimental setup:

BL14.1:
- Photon energy range: 5.5-16 keV (wavelength: 0.77502.25 A)
- Photon flux: 1.8x10¹¹ Phot/sec x 100 mA at sample position
 (0.1-1 sec exposure time per frame)
- PILATUS 6M detector, 141 mm-680 mm max. distance from the sample
- Microdiffractometer (MD2) with Mini-kappa goniometer
- Automatic sample changer (CATS), 90 sample storage capacity
 (SPINE-Pin & EMBL sample magazine compatibility)
- User defined beam shaping from 50 µm-100 µm diameter possible
- In situ crystal-screening using 96-well plates
- 32-core XEON-CPU server, with 10GB uplink to Pilatus 6M
- Data collection control via MXCuBE2
- Common MX-software installed including EDNA, XDS, iMOSFLM, CCP4,
 Phenix, SHELXC-D-E, etc.
- Automated data processing using XDSAPP
- Remotely controlled cryo-shutter for crystal annealing
- AMPTEK-XRF detector and XFEPLOT software available

We are also offering the hard- and software environment for
carrying out UV-RIP experiments at BL14.1. For further information,
please visit:
https://www.helmholtz-berlin.de/forschung/oe/np/gmx/ancillary-facilities/uvrip_en.html

BL14.2:
- Photon energy range: 5.5-16 keV (wavelength: 0.77502.25 A)
- Photon flux: 1.6x10¹¹ Phot/sec x 100 mA at sample position
 (0.05-1 sec exposure time per frame)
- PILATUS 3S-2M detector with 1000 µm Si sensor thickness,
 85 mm - 800 mm distance from the sample (a special mode with
 55 mm distance is also available upon request)
- Nanodiffractometer with fast air-bearing axis and on-axis sample
 microscope
- User defined beam shaping from 30 µm-150 µm diameter possible
- Data collection control via MXCuBE2
- G-ROB sample changer for SPINE and UNIPUCK support
- 60-core XEON-CPU server, with fibre channel SAN up-link data
 processing environment
- Common MX-software installed including EDNA, XDS, iMOSFLM, CCP4,
 Phenix, SHELXC-D-E, etc.
- Automated data processing using XDSAPP
- AMPTEK-XRF detector and XFEPLOT software available
- UV-Microsprectrophotometer offline setup available

If you need atomic resolution or better, BL14.2 is the beamline
of choice for you.

BL14.3:
- Fixed photon energy: 13.8 keV (wavelength: 0.89 A)
- Photon flux: 1.6x10exp10 Phot/sec x 100mA at sample position
 (3-20 sec exposure time per frame)
- Rayonix MX-225 X-ray detector, 54 mm-450 mm distance from the
 sample
- MD2S microdiffractometer with mini-kappa goniometer
- In situ crystal-screening using 96-well plates
- RT data collection
- 60-core XEON-CPU server, with fibre channel SAN up-link data
 processing environment
- Data collection control via MXCuBE2
- Common MX software installed including EDNA, XDS, iMOSFLM, CCP4,
 Phenix, SHELXC-D-E, etc.
- Automated data processing using XDSAPP
- Remotely controlled cryo-shutter for crystal annealing
- REX rapid nozzle exchanger
- HC-Lab dehydration device installed (please specify HC-Lab-beamtime
 in your proposal if needed)
- AMPTEK-XRF detector and XFEPLOT software available

Other facilities:
- Ultra high performance stereo microscope Leica M205A, 20-255x zoom,
 8 Mpixel CCD-camera
- Pressure chamber for noble gas derivatization (Xe, Kr available
 upon request)

S1-biolab facilities (separate registration required):
- Protein production and purification (AEKTA)
- nL 96-well crystallization plate formulation and storage at
 5°C and 20°C
- Biophysical characterization with real time PCR (thermofluor assay)
- Contactless compound pipetting using ATS

The HZB-MX group is also providing expert assistance as well as
access to a library of fragments for carrying out crystallographic
fragment-screening experiments. For more information please see