Protein Crystallography

Main equipment

Crystallization robot – Honeybee963®:
The Honeybee963® (Digilab) robot is a bench-top system for the automation and miniaturization of vapor diffusion in sitting-drop protein crystallization experiments. Proprietary Cartesian synQUAD® dispensers couple high-speed micro-solenoid valves with high resolution syringe pumps, dispensing volumes down to 100 nL. We currently use 200-300 nL nanodrops to maximize precision and crystallizability. The 96-needle arm allows for very fast dispensing of reservoir solutions on a 96-well setup. Three independent protein synQUAD® needles then proceed to dispense up to three different proteins, variable volumes are defined using the robot’s software. Automation enables the assay of typically hundreds of different potential crystallogenesis conditions in a matter of minutes, allowing to greatly increase the search space, and thus the probability of finding hits.

Automatic setup and optimization of screen assays – Alchemist DT®:
The Alchemist DT ® (Rigaku) is a bench-top liquid handling robot for the screen production and optimization of crystallization conditions. It provides consistent, precise and accurate liquid dispensing in a volume range of 1 μl to 10 ml into SBS, Linbro ® and Nextal® footprint plates. Due to its technology, elimination of tubing means no waste and removes the possibility of cross-contamination.
CrystalTrak™, the integrated software package, is designed specifically for protein crystallography. Once the screen is designed, CrystalTrak™ automatically calculates the recipe and defines the necessary stock solutions for use with the Alchemist. 26 different stock solutions can be stored on the deck at one time. Stock management tools and barcode tracking ensure that the correct stock solutions and necessary volumes of solutions are available on the deck before any plate generation begins.

X ray generator – Rigaku MicroMax-007HF®:
Micromax007-HF® (Rigaku) is an X-ray generator with a 0.07 mm diameter effective focal size at the source. Equipped with a Cu rotating anode, it provides an output of 1200 W and a brightness of 31 kW/mm2. In combination with the installed optics (Varimax-HF®, Rigaku) which consists of confocal multilayer mirrors, the resulting X rays focused on the crystalline sample are ultra-bright, and can be used effectively for various measurement purposes. We can solve structures with atoms that scatter anomalously at 1.5418Å wavelength (S, I, Cs, lanthanides, among the most used). Molecular replacement problems can be tackled, as well as data collection for high-resolution refinement (including ligands, inhibitors, drugs, point-mutation protein variants, etc).

Image plate area detector – MAR345®:
The MAR345® (Mar Research) detector installed on a MAR345dtb®table is an image plate detector that enables us to collect data up to 1.2Å resolution on our geometric setup (taking full advantage of the 2θ angle). It is a single Φ-angle oscillation setup, equipped with a convenient χ-motor that facilitates crystal mounting under cryogenic conditions. Read-out cycles range from 108 to 34 seconds, depending on pixel size and effectively scanned plate diameter.
The read-out system of the Mar345 is unique in its use of a single high performance 85mW laser which delivers more than 0.8 µJ/pixel at the plate. This ensures that an extremely high percentage (>95%) of trapped F-centers are transformed into photostimulated luminiscence.