We offer the possibility to test more than 500 crystallisation conditions, thanks to the use of a nano-drop dispenser robot (Honeybee) with a wide choice of commercial screening kits (primarily the Nextal series by Quiagen). These crystallisation tests can be performed in the presence of ligands or additives (for example detergents). The technique used for these assays is the sitting drop method, which requires at least 0.5 microL of protein solution per 96-well plate. If these initial tests result in (micro-) crystals, screening on the crystallogenesis conditions can be performed in order to improve the quality and size of the crystals.
Cristallography
The platform for crystallography is located on site of the Station Biologique de Roscoff. It is composed of two parts: i) Crystallogenesis ii) Diffraction - 3D Structure Determination & Refinement.
Expertise
The personnel of the platform accompanies and guides any structural project that uses the approach of protein crystallography. More specifically the personnel can :
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Assure and accompany the access to the machines (crystallization robot & diffractometer, as well as access to synchrotron radiation)
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Evaluate the feasibility (time needed, cost, novelty, difficulty..) of presented projects
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Follow and accompany the projects along all steps of the approach
Principal Equipments
> A unit for production and detection of X-rays : rotating anode NONIUS/BRUKER.
Diffractometer with microfocus, equipped with a four-circle goniometer and a bi-dimensional CCD detector.
Cryocooling facility and installation to freeze, store and collect the data at 100 K
> Nano-drop dispensing robot for crystallogenesis
A nano-drop dispensing robot of the type ‘Honeybee’ from Protein Solutions, for the setup of crystallization trials in 96-well format.
Rooms/storage to setup trials at 18°C and 4°C.
> Access to a informatics server and dedicated software
1 calculation cluster with 6 processors (Linux); network of 6 PC (linux) with graphical devices for data treatment and analysis. Support for the use of all classical software suites used in protein crystallography for interactive construction, visualization and analysis of 3D biomolecule structures.
Prestations
Structure determination
Prestation :
Diffraction, determination and refinement of 3D structures
Services :
Sites :
If the crystallogenesis assays succeed in obtaining crystals with a diffractive ability of at least 3 Angstroms on the synchrotron (ESRF in Grenoble), determination of the structure will be initiated. If the bioinformatic analysis reveals structural homologues of the crystallised protein, resolution of the structure will be determined through molecular replacement. Otherwise, heavy atom derivatives can be prepared (selenomethionine, selenocysteine, etc.), in order to use the different isomorphous replacement methods.
- crystallogenesis of biological macromolecules,
- diffraction data collection on-site and from synchrotron sources,
- determination and refinement of 3D structures,
- molecular modelling: homologue proteins, protein-protein interactions (etc.),
- digital tools for analysis of structure/function relationships,
- development of potential applications.