Granada Crystallization Facility (GCF) - 11.22.16
The Granada Crystallization Facility (GCF) is a multiuser facility designed to conduct crystallization experiments of biological macromolecules in microgravity using a counterdiffusion technique inside capillaries. The counterdiffusion technique is implemented in the Granada Crystallization Boxes (GCBs). Science Results for Everyone
Information Pending Facility Details
Juan Manuel Garcia-Ruiz, Ph.D., University of Granada, Granada, Spain
Fermin Otalora Munoz, University of Granada, Granada, Spain
Luis Antonio Gonzalez Ramirez, University of Granada, Granada, Spain
Jose Antonio Gavira, Laboratorio de Estudios Crystallograficos, University of Granada, Granada, Spain
Triana Science and Technology, Granada, Spain
NTE, Barcelona, Spain
Sponsoring Space Agency
European Space Agency (ESA)
ISS Expedition Duration
August 2001 - December 2001; June 2002 - December 2002; November 2002 - May 2003; April 2003 - April 2005; October 2005 - April 2006; September 2006 - October 2008
The Grenada Crystallization Facility (GCF) has been used in several European Space Agency (ESA) and Japan Aerospace Exploration Agency (JAXA) GCF investigations on the International Space Station.
- The Granada Crystallization Facility (GCF) was designed to crystallize macromolecules in space at a very low cost to ensure maximum success.
- The GCF is autonomous; it does not require crew time during operations, and it is a passive device (it does not need electric power to work). The GCF can hold 300 crystallization experiments with a volume of 1 L and a mass of approximately 1 kg.
- The GCF works by counterdiffusion to minimize the effects of surface-driven convection and g-jitter on the experiments. In addition to its low cost, the GCF has the following advantages:
- It works under diffusion-controlled mass transport.
- The counterdiffusion technique automatically searches for the optimal crystallization conditions.
- The volume of protein required for the experiment is minimized.
Counterdiffusion crystallization experiments have been performed in the GCB. The bottom of the box contains a layer of buffered agarose gel at a 0.5 percent w/v concentration. Capillaries with diameters ranging from 0.2 to 1.5 mm are used as protein chambers. Once the gel is set, the capillaries are filled with the protein, inserted through the hole of the capillary holder, and inserted at a given depth into the gelled buffer layer. This depth represents the effective width of the buffer layer for passive activation and will be calculated as a function of the time elapsed between the time the reactor is filled and the time it is placed in its final location in the International Space Station (ISS). Finally the salt solution is poured on top of the buffer gel.
The GCB can be used as a passive reactor without moving parts or crew manipulation for microgravity experiments. The method is simple. The crystallization reactors must be supplied for integration into the spacecraft several hours before launch (the waiting time for launch). After takeoff, it takes time to reach orbit around the Earth (about 8.5 minutes) and, in the case of the ISS, to be locked to the ISS and place the crystallization reactor in the proper location (the waiting time for orbit). Taking into account these considerations, the capillaries are punching into the gel with a depth x such that x > (Dt)1/2, where "D" is the diffusion coefficient of the precipitating agent and "t" is the waiting time for launch plus the waiting time for orbit. As a result, during the waiting times for launch and orbit, the precipitating agent is diffusing across the gel, and convection is precluded. Once the GCB is orbiting the Earth, the precipitating agent will reach the protein solution filling the capillaries, and crystallization will take place in the gel-free solution under diffusion-controlled mass transport. Therefore, the experiment in the GCF can be considered totally passive, with no activation and/or manipulation by crewmembers required. Protein crystallization experiments in the International Space Station were performed using Granada Crystallization Boxes (GCBs), each of which accommodates up to six capillary counterdiffusion experiments.
The Granada Crystallization Box (GCB) is used to perform counterdiffusion experiments. The GCB consists of three elements made of polystyrene: a box into which the gel is introduced, a capillary holder to hold capillaries, and a cover. The buffer layer is applied at the bottom of the reservoir after the capillary holder is inserted. Once the gel is set, the capillaries are filled with the protein, inserted through the hole of the capillary holder, and inserted to a given depth into the gelled buffer layer. The salt solution is set on top of the buffer gel.
The GCF is a passive device. The hardware requires no crew time on orbit except the time it takes to place and remove the GCF. ^ back to top
- The Grenada Crystallization Box (GCB) samples were preloaded in the Grenada Crystallization Facility (GCF) at the launch site 2 days before takeoff for delivery to the International Space Station (ISS).
- After arriving at the ISS, the GCF is installed in either the Russian Kubik incubator or the U.S. Commercial Generic Bioprocessing Apparatus (CGBA). The GCF remains in place after the end of the mission.
- The GCF is returned to Earth. After landing, the experiments are returned to the investigators for analysis of the results.
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Protein crystallization experiments on the International Space Station (ISS)were performed on the Andromede and Odissea missions using 23 Grenada Crystallization Boxes (GCBs). Each GCB accommodates as many as six capillary counterdiffusion experiments.
The Grenada Crystallization Facility (GCF) is an aluminium container with internal dimensions of 12 x 12 x 8 cm. The GCFs for the GCBs were designed, built, and tested to ensure that they were capable of withstanding launch and reentry maneuvers and met ISS safety requirements.
The preliminary analysis of the data sets collected with synchrotron radiation shows that the crystals grown using the counterdiffusion technique share excellent global indicators of x-ray data quality, with no obvious difference between crystals grown in reduced-convection conditions in space and crystals grown in convection-free conditions on the ground.
Maes D, Decanniere K, Zegers I, Vanhee C, Sleutel M, Willaert R, Van de Weerdt C, Martial J, Declercq J, Evrard C, Otalora Munoz F, Garcia-Ruiz JM. Protein crystallisation under microgravity conditions: What did we learn on TIM crystallisation from the Soyuz missions?. Microgravity Science and Technology. 2007; XIX-5/6: 90-94.
Garcia-Ruiz JM, Gonzalez Ramirez LA, Gavira JA, Otalora Munoz F. Granada Crystallisation Box: a new device for protein crystallisation by counter-diffusion techniques. Acta Crystallographica Section D: Biological Crystallography. 2002 September 26; 58(10): 1638-1642. DOI: 10.1107/S0907444902014464.
Ground Based Results Publications
Garcia-Ruiz JM. Counterdiffusion methods for macromolecular crystallization. Methods in Enzymology. 2003; 368: 130-154. DOI: 10.1016/S0076-6879(03)68008-0. PMID: 14674272.
Zegers I, Carotenuto L, Evrard C, Garcia-Ruiz JM, De Gieter P, Gonzales-Ramires L, Istasse E, Legros J, Martial J, Minetti C, Otalora Munoz F, Queeckers P, Cedric S, Van de Weerdt C, Willaert R, Wyns L, Yourassowsky C, Dubois F. Counterdiffusion protein crystallisation in microgravity and its observation with PromISS (Protein Microscope for the International Space Station). Microgravity Science and Technology. 2006; 18-3/4: 165-169.
Tanaka H, Inaka K, Sugiyama S, Takahashi S, Sano S, Sato M, Yoshitomi S. A simplified counter diffusion method combined with a 1D simulation program for optimizing crystallization conditions. Journal of Synchrotron Radiation. 2004; 11: 45-48.
Garcia-Ruiz JM. Nucleation of protein crystals. Journal of Structural Biology. 2003; 142: 22-31.
Evrard C, Maes D, Zegers I, Declercq J, Vanhee C, Martial J, Wyns L, Van de Weerdt C. TIM Crystals Grown by Capillary Counterdiffusion: Statistical Evidence of Quality Improvement in Microgravity. Crystal Growth and Design. 2007 November; 7(11): 2161-2166. DOI: 10.1021/cg700687t.
Ng JD, Gavira JA, Garcia-Ruiz JM. Protein crystallization by capillary counterdiffusion for applied crystallographic structure determination. Journal of Structural Biology. 2003; 142: 218-231.
- Granada Crystallization Facility in Odissea
- The Grenada Crystallization Facility on the Andromede Mission
Granada Crystallization Box hardware. Image courtesy of Triana Science and Technology, Granada, Spain.
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NASA Image: ISS003E5331 - Expedition Three Flight Engineer Vladimir N. Dezhurov unpacks a bag with the Granada Crystallization Facility in the Zvezda service module during the Andromede mission.
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NASA Image: ISS003E5394 - The Expedition 3 crewmembers, Flight Engineer Mikhail Tyurin (left), Mission Commander Frank L. Culbertson (center), and Flight Engineer Vladimir Dezhurov (right), assemble for a group photo in the Zvezda service module of the ISS. In the background, the Granada Crystallization Facility is visible in the orange container.
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