X-ray structure determination of the glycine cleavage system protein H of Mycobacterium tuberculosis using an inverse Compton synchrotron X-ray source
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Structural genomics discovery projects require ready access to both X-ray diffraction and NMR spectroscopy which support the collection of experimental data needed to solve large numbers of novel protein structures. The most productive X-ray crystal structure determination laboratories make extensive use of tunable synchrotron X-ray light to solve novel structures by anomalous diffraction methods. This requires that frozen cryo-protected crystals be shipped to large multi acre synchrotron facilities for data collection. In this paper we report on the development and use of the first laboratory-scale synchrotron light source capable of performing many of the state-of-the-art synchrotron applications in X-ray science. This Compact Light Source is a first-in-class device that uses inverse Compton scattering to generate X-rays of sufficient flux, tunable wavelength and beam size to allow high-resolution X-ray diffraction data collection from protein crystals. We report on benchmarking tests of X-ray diffraction data collection with hen egg white lysozyme, and the successful high-resolution X-ray structure determination of the Glycine cleavage system protein H from Mycobacterium tuberculosis using diffraction data collected with the Compact Light Source X-ray beam.
KeywordsCompact light source X-ray crystallography Inverse Compton
We gratefully acknowledge NIH funding for the developments included in this paper. The prototype CLS was funded by the NIGMS under grant R44-GM66511. The X-ray station and optics development was funded by the NIGMS and NCRR under grant R44-GM074437. The development of the Beta CLS was funded by the NIGMS and NCRR which have co-sponsored the Accelerated Technologies Center for Gene to 3D Structure (www.atcg3d.org), a PSI-2 Specialized Center, under Grant U54 GM074961. The production of MytuGCSPH protein and crystals was funded by NIAID under Federal Contract No. HHSN272200700057C which supports the Seattle Structural Genomics Center for Infectious Disease (www.SSGCID.org). Special thanks to Shellie Dieterich, Becky Poplawski, and Jeff Christensen at Emerald BioStructures for their support in molecular biology and protein crystallization. The authors also wish to thank the organizing committee of the PSI organized Enabling Technologies Meetings held each spring on the NIH campus. LTI collaborators wish to thank the staff at LTI, Michael Blum, Jens Als-Nielsen, Anette Jensen and Bill Weis.
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