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July 7, 2004 NSLS 2004 Annual Users’ Meeting WorkshopCrystallization: Focus on Membrane ProteinsThe Crystallization workshop organized in conjunction with the 2004 National Synchrotron Light Source Annual Users’ Meeting focused this year on crystallization techniques available to membrane proteins. This two day workshop covered different crystallization methods from the most standard vapor diffusion to more advanced techniques. A morning of introductory talks was followed by five practicals, of 2 hours each, where participants had the opportunity to carry out different crystallization methods in the Laboratory. The main purpose of this workshop was to bring the practical aspect of the crystallization methods to the participants; the exchange of experiences and ideas being its essence.
Naomi Chayen, Imperial College London, Tackling the bottleneck of protein crystallization: Practical techniques with a difference introduced the phase diagram and discussed the several different crystallization techniques developed for soluble proteins and the necessary modifications for crystallization of membrane proteins. The laboratory practicals included setting up of microbatch and vapour diffusion trials using oils as a tool to aid crystal growth. Participants had opportunity to try several different crystallization configurations. Peter Nollert, deCODE Genetics, Miniaturization of the cubic phase, introduced the cubic lipid phase method and discussed the difficulties of membrane protein crystallization. The lipid cubic phase method was demonstrated in the Laboratory and participants were able to set up crystallization trials for bacteriorhodopsin. The application to soluble proteins was also introduced. A number of participants tried the method with their own proteins with success. Petra Fromme, Arizona State University, Overcoming the crystallization problems of PSI and PSII, discussed the importance of the phase diagram to the crystallization of the PSI and PSII proteins. The contribution of the dialysis method was introduced and participants were able to assemble their own crystallization reactors. Most experiments lead to the crystallization of PSI within 24 hours and by the end of the workshop participants were able to harvest some of the crystals. Marie Claude Marchand, NEXTAL Biotechnologies, The Vapour Diffusion Method, discussed the vapour diffusion methods, seeding, derivatization, and optimization of the phase diagram. Some participants were able to screen for the best crystallization conditions of their own protein Ana Belen Moradela Merlo, University of Granada, Protein Crystallization by the Counter diffusion, introduced the counter diffusion method. Participants were able to set up crystallization trials of soluble proteins in the “Granada Box”. Possible applications to membrane proteins were discussed. The demand for the course was overwhelming. It was oversubscribed by over 100%. At the end of the workshop students were asked to evaluate the course and suggest changes and additions; two thirds responded to the survey. Overall the course was rated as extremely useful as reflected in one of the answers to the survey … “I learned a great deal about crystallization methods. Most importantly I now understand the more practical aspects of each method that I was not aware of. These ideas cannot be gained from other formats”. All participants who answered the survey were of the opinion that the workshop should be offered again. ACKNOWLEDGEMENTS FOR MORE INFORMATION Naomi Chayen |
The National Synchrotron Light Source at Brookhaven National Laboratory in New York, is a national user research
facility funded by the U.S. Department of Energy's Office of Basic Energy Science. The NSLS operates two electron
storage rings: an X-Ray ring and a Vacuum UltraViolet (VUV) ring which provide intense light spanning the electromagnetic
spectrum from the infrared through x-rays. Each year over 2300 scientists from universities, industries and government
labs perform research at the NSLS.
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