Wide Angle X-Ray Scattering
Lee Makowski, Principle Investigator
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4HHB: The crystal structure of human deoxyhaemoglobin at 1.74 A resolution |
Wide angle x-ray scattering from proteins in solution produces data that contains information relevant to determination of protein fold. But at relevant scattering angles these data are weak, and the degree to which they might be used to categorize the fold of a protein is unknown. Our work at BioCAT (sector 18 of the Advanced Photon Source) has demonstrated the capability of collecting scattering data from proteins in solution to spacings of 2.2 Ǻ (q=2.8 A-1), and the potential for the use of this data in structural characterization of proteins. Feasibility studies performed at the BioCAT undulator beam line at the APS indicate that WAXS studies of protein solutions have significant potential for closing the gap between target and fold designation that presently exists in the structural genomics pipeline. This project seeks to optimize and enhance the collection and analysis of solution scattering data and to determine the extent to which it is possible to accurately determine the fold of a protein from this data, as well as study real-time changes in conformation in response to ligand binding, denaturants, etc.
- WAXs Distinguishes Ligand Affinity in Receptor Binding Events
- MAD MAX as a Novel Tool for the Calculation of Internal Protein Movement
- Characterization of Protein Breathing Using Coordinated WAXS and NSE
Our group has designed an apparatus (below) for the initial studies using WAXS of protein solutions with minimal air gaps. The experimental set-up includes a thermostatistacally controlled copper block (using fluid flow from a re-circulation bath) which encloses the quartz capillary through which the protein solution is pulled via a programmable peristaltic pump. There is also coupled control of protein flow through the sample and shutter opening so that the protein needs to start flowing immediatly before the shutter opens, and stop flowing immediatly after the shutter closes.
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Photos courtesy of D.J. Rodi and R.F. Fischetti |
Use of the Advanced Photon Source was supported by the U.S. Department of Energy, Basic Energy Sciences, Office of Science. BioCAT is a National Institutes of Health-supported Research Center RR-08630.
