TY - JOUR
T1 - Toward structure determination using membrane-protein nanocrystals and microcrystals
AU - Hunter, Mark S.
AU - Fromme, Petra
N1 - Funding Information:
The Advanced Light Source is supported by the Director, Office of Science, Office of Basic Energy Sciences, of the US Department of Energy under Contract No. DE-AC02-05CH11231.
Funding Information:
We acknowledge support from the Helmholtz Association ; the Max Planck Society , for funding the development and operation of the CAMP instrument within the ASG at CFEL; DOE, through the PULSE Institute at the SLAC National Accelerator Laboratory, and by the Lawrence Livermore National Laboratory under Contract DE-AC52-07NA27344; the US National Science Foundation (awards 0417142 and MCB-1021557); the US National Institutes of Health (awards 1R01GM095583-01 (ROADMAP) and 1U54GM094625-01 (PSI:Biology)); the Joachim Herz Stiftung , the Swedish Research Council ; the Swedish Foundation for International Cooperation in Research and Higher Education , Stiftelsen Olle Engkvist Byggmastare .
Funding Information:
This work was supported by NSF award IDBR 0555845, the Center for Biophotonics Science and Technology (University of California at Davis) , the Lawrence Berkeley National Laboratory Seaborg Fellowship award, by the US Department of Energy through the PULSE Institute at the SLAC National Accelerator Laboratory , and by Lawrence Livermore National Laboratory under Contract DE-AC52-07NA27344 , and the Joachim Herz Stiftung.
PY - 2011/12
Y1 - 2011/12
N2 - Membrane proteins are very important for all living cells, being involved in respiration, photosynthesis, cellular uptake and signal transduction, amongst other vital functions. However, less than 300 unique membrane protein structures have been determined to date, often due to difficulties associated with the growth of sufficiently large and well-ordered crystals. This work has been focused on showing the first proof of concept for using membrane protein nanocrystals and microcrystals for high-resolution structure determination. Upon determining that crystals of the membrane protein Photosystem I, which is the largest and most complex membrane protein crystallized to date, exist with only 100. unit cells with sizes of less than 200. nm on an edge, work was done to develop a technique that could exploit the growth of the Photosystem I nanocrystals and microcrystals. Femtosecond X-ray protein nanocrystallography was developed for use at the first high-energy X-ray free electron laser, the LCLS at SLAC National Accelerator Laboratory, in which a liquid jet brought fully-hydrated Photosystem I nanocrystals into the interaction region of the pulsed X-ray source. Diffraction patterns were recorded from millions of individual PSI nanocrystals and data from thousands of different, randomly oriented crystallites were integrated using Monte Carlo integration of the peak intensities. The short pulses (~70. fs) provided by the LCLS allowed the possibility to collect the diffraction data before the onset of radiation damage, exploiting the diffract-before-destroy principle. During the initial experiments at the AMO beamline using 6.9-Å wavelength, Bragg peaks were recorded to 8.5-Å resolution, and an electron-density map was determined that did not show any effects of X-ray-induced radiation damage [94]. Many additional techniques still need to be developed to explore the femtosecond nanocrystallography technique for experimental phasing and time-resolved X-ray crystallography experiments. The first proof-of-principle results for the femtosecond nanocrystallography technique indicate the incredible potential of the technique to offer a new route to the structure determination of membrane proteins.
AB - Membrane proteins are very important for all living cells, being involved in respiration, photosynthesis, cellular uptake and signal transduction, amongst other vital functions. However, less than 300 unique membrane protein structures have been determined to date, often due to difficulties associated with the growth of sufficiently large and well-ordered crystals. This work has been focused on showing the first proof of concept for using membrane protein nanocrystals and microcrystals for high-resolution structure determination. Upon determining that crystals of the membrane protein Photosystem I, which is the largest and most complex membrane protein crystallized to date, exist with only 100. unit cells with sizes of less than 200. nm on an edge, work was done to develop a technique that could exploit the growth of the Photosystem I nanocrystals and microcrystals. Femtosecond X-ray protein nanocrystallography was developed for use at the first high-energy X-ray free electron laser, the LCLS at SLAC National Accelerator Laboratory, in which a liquid jet brought fully-hydrated Photosystem I nanocrystals into the interaction region of the pulsed X-ray source. Diffraction patterns were recorded from millions of individual PSI nanocrystals and data from thousands of different, randomly oriented crystallites were integrated using Monte Carlo integration of the peak intensities. The short pulses (~70. fs) provided by the LCLS allowed the possibility to collect the diffraction data before the onset of radiation damage, exploiting the diffract-before-destroy principle. During the initial experiments at the AMO beamline using 6.9-Å wavelength, Bragg peaks were recorded to 8.5-Å resolution, and an electron-density map was determined that did not show any effects of X-ray-induced radiation damage [94]. Many additional techniques still need to be developed to explore the femtosecond nanocrystallography technique for experimental phasing and time-resolved X-ray crystallography experiments. The first proof-of-principle results for the femtosecond nanocrystallography technique indicate the incredible potential of the technique to offer a new route to the structure determination of membrane proteins.
KW - Femtosecond nanocrystallography
KW - Membrane proteins
KW - Protein nanocrystals
KW - Structure determination
KW - X-ray crystallography
KW - XFEL
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U2 - 10.1016/j.ymeth.2011.12.006
DO - 10.1016/j.ymeth.2011.12.006
M3 - Review article
C2 - 22197730
AN - SCOPUS:84855994087
SN - 1046-2023
VL - 55
SP - 387
EP - 404
JO - Methods
JF - Methods
IS - 4
ER -