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Title: Development of the XFP beamline for x-ray footprinting at NSLS-II

Abstract

For over a decade, synchrotron-based footprinting studies at the NSLS X28C beamline have provided unique insights and approaches for examining the solution-state structures of large macromolecular assemblies, membrane proteins, and soluble proteins, for time-resolved studies of macromolecular dynamics, and most recently for in vivo studies of RNA-protein complexes. The transition from NSLS to NSLS-II has provided the opportunity to create an upgraded facility for the study of increasingly complex systems; progress on the development of the XFP (X-ray Footprinting for In Vitro and In Vivo Structural Studies of Biological Macromolecules) beamline at NSLS-II is presented here. The XFP beamline will utilize a focused 3-pole wiggler source to deliver a high flux density x-ray beam, where dynamics can be studied on the microsecond to millisecond timescales appropriate for probing biological macromolecules while minimizing sample perturbation. The beamline optics and diagnostics enable adaptation of the beam size and shape to accommodate a variety of sample morphologies with accurate measurement of the incident beam, and the upgrades in sample handling and environment control will allow study of highly sensitive or unstable samples. The XFP beamline is expected to enhance relevant flux densities more than an order of magnitude from that previously available atmore » X28C, allowing static and time-resolved structural analysis of highly complex samples that have previously pushed the boundaries of x-ray footprinting technology. XFP, located at NSLS-II 17-BM, is anticipated to become available for users in 2016.« less

Authors:
; ; ; ;  [1];  [2]
  1. Case Center for Synchrotron Biosciences, Center for Proteomics and Bioinformatics, Case Western Reserve University, Cleveland, OH (United States)
  2. Brookhaven National Laboratory, Upton, NY (United States)
Publication Date:
OSTI Identifier:
22608342
Resource Type:
Journal Article
Resource Relation:
Journal Name: AIP Conference Proceedings; Journal Volume: 1741; Journal Issue: 1; Conference: SRI2015: 12. international conference on synchrotron radiation instrumentation, New York, NY (United States), 6-10 Jul 2015; Other Information: (c) 2016 Author(s); Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
43 PARTICLE ACCELERATORS; BEAMS; DISTURBANCES; FLUX DENSITY; MEMBRANE PROTEINS; MEMBRANES; NSLS; PERTURBATION THEORY; RNA; SYNCHROTRONS; TIME RESOLUTION; X RADIATION

Citation Formats

Bohon, Jen, E-mail: jbohon@bnl.gov, Sullivan, Michael, Abel, Don, Toomey, John, Chance, Mark R., E-mail: mark.chance@case.edu, and Dvorak, Joseph. Development of the XFP beamline for x-ray footprinting at NSLS-II. United States: N. p., 2016. Web. doi:10.1063/1.4952828.
Bohon, Jen, E-mail: jbohon@bnl.gov, Sullivan, Michael, Abel, Don, Toomey, John, Chance, Mark R., E-mail: mark.chance@case.edu, & Dvorak, Joseph. Development of the XFP beamline for x-ray footprinting at NSLS-II. United States. doi:10.1063/1.4952828.
Bohon, Jen, E-mail: jbohon@bnl.gov, Sullivan, Michael, Abel, Don, Toomey, John, Chance, Mark R., E-mail: mark.chance@case.edu, and Dvorak, Joseph. 2016. "Development of the XFP beamline for x-ray footprinting at NSLS-II". United States. doi:10.1063/1.4952828.
@article{osti_22608342,
title = {Development of the XFP beamline for x-ray footprinting at NSLS-II},
author = {Bohon, Jen, E-mail: jbohon@bnl.gov and Sullivan, Michael and Abel, Don and Toomey, John and Chance, Mark R., E-mail: mark.chance@case.edu and Dvorak, Joseph},
abstractNote = {For over a decade, synchrotron-based footprinting studies at the NSLS X28C beamline have provided unique insights and approaches for examining the solution-state structures of large macromolecular assemblies, membrane proteins, and soluble proteins, for time-resolved studies of macromolecular dynamics, and most recently for in vivo studies of RNA-protein complexes. The transition from NSLS to NSLS-II has provided the opportunity to create an upgraded facility for the study of increasingly complex systems; progress on the development of the XFP (X-ray Footprinting for In Vitro and In Vivo Structural Studies of Biological Macromolecules) beamline at NSLS-II is presented here. The XFP beamline will utilize a focused 3-pole wiggler source to deliver a high flux density x-ray beam, where dynamics can be studied on the microsecond to millisecond timescales appropriate for probing biological macromolecules while minimizing sample perturbation. The beamline optics and diagnostics enable adaptation of the beam size and shape to accommodate a variety of sample morphologies with accurate measurement of the incident beam, and the upgrades in sample handling and environment control will allow study of highly sensitive or unstable samples. The XFP beamline is expected to enhance relevant flux densities more than an order of magnitude from that previously available at X28C, allowing static and time-resolved structural analysis of highly complex samples that have previously pushed the boundaries of x-ray footprinting technology. XFP, located at NSLS-II 17-BM, is anticipated to become available for users in 2016.},
doi = {10.1063/1.4952828},
journal = {AIP Conference Proceedings},
number = 1,
volume = 1741,
place = {United States},
year = 2016,
month = 7
}
  • Structural mapping of proteins and nucleic acids with high resolution in solution is of critical importance for understanding their biological function. A wide range of footprinting technologies have been developed over the last ten years to address this need. Beamline X28C, a white-beam X-ray source at the National Synchrotron Light Source of Brookhaven National Laboratory, functions as a platform for synchrotron footprinting research and further technology development in this growing field. An expanding set of user groups utilize this national resource funded by the National Institute of Biomedical Imaging and Bioengineering of the National Institutes of Health. The facility ismore » operated by the Center for Synchrotron Biosciences and the Center for Proteomics of Case Western Reserve University. The facility includes instrumentation suitable for conducting both steady-state and millisecond time-resolved footprinting experiments based on the production of hydroxyl radicals by X-rays. Footprinting studies of nucleic acids are routinely conducted with X-ray exposures of tens of milliseconds, which include studies of nucleic acid folding and their interactions with proteins. This technology can also be used to study protein structure and dynamics in solution as well as protein-protein interactions in large macromolecular complexes. This article provides an overview of the X28C beamline technology and defines protocols for its adoption at other synchrotron facilities. Lastly, several examples of published results provide illustrations of the kinds of experiments likely to be successful using these approaches.« less
  • X-ray footprinting employs intense X-rays produced by synchrotron radiation to generate hydroxyl radicals in solution on microseconds-milliseconds timescales. These hydroxyls radicals undergo stable reaction with solvent accessible sites of macromolecule and produce covalent modifications, which are appropriate to probing macromolecule dynamics under physiological condition. For nucleic acids, one analyzes the pattern of fragments after X-ray exposure by gel electrophoresis; the protected sections that are not cleaved yield a 'footprint'. For proteins, the exposed samples are digested with proteases and analyzed by liquid chromatography- and tandem-mass spectrometry to determine the extent and sites of modification. The data provide detailed structural informationmore » to map tertiary contacts of macromolecular interactions, which can subsequently be used as constraints for molecular modeling to generate high-resolution structures. This method is unique in providing 'local' structural information in solution for gaining insight into dynamic processes involving, large RNA-protein and protein-protein assemblies on biologically relevant timescales. The method also can uniquely probe the 'local' structure of large complexes poised at equilibrium for functional states of interest, and has been extended to in vivo studies. Beamline X28C is located at the National Synchrotron Light Source of Brookhaven National Laboratory. An expanding set of user groups utilize this national resource funded by the National Institute of Biomedical Imaging and Bioengineering of the National Institutes of Health. The facility is operated by the Center for Synchrotron Biosciences and the Center for Proteomics and Bioinformatics of Case Western Reserve University. The facility supports both onsite and offsite user access. Beam time is allocated online through peer reviewed user proposal system. Examples of recent research projects are provided.« less
  • The U7A bending magnet toroidal grating monochromator (TGM) beamline has been converted into a spherical grating monochromator (SGM) type, without constructing any new vacuum components. That is, the mirror, slit, and grating chambers have been recycled. The inherited optical arrangement and the recycling concept prevented the use of {open_quote}{open_quote}standard{close_quote}{close_quote} SGM Kirkpatrick-Baez collecting and focusing mirrors at the front end. We show that this modification is not a serious detriment to the optical properties of the resulting beamline. Specifically, the resolution is completely unaffected and the throughput is approximately 2/3 of the {open_quote}{open_quote}standard{close_quote}{close_quote} SGM value. The flux and photon energy resolutionmore » of the U7A beamline throughout its 200{endash}1000 eV operating photon energy range will be presented. Three end stations are being constructed for this beamline, featuring soft x-ray absorption and photoemission chambers optimized for operation in the carbon K-, oxygen K-, and transition metal L-edge ranges. Two of these end stations will be preceded by refocusing mirrors that will focus the soft x-rays to {lt}1 mm spot size onto their samples. {copyright} {ital 1996 American Institute of Physics.}« less
  • A newly designed monochromator assembly for the State University of New York (SUNY) X3 beamline at the National Synchrotron Light Source (NSLS) consists of two monochromators enclosed in a single vessel, positioned at 7813 mm from the bending magnet source. It allows delivery of monochromatic radiation into two independent end stations. The first unit contains two crystals reflecting in the vertical plane, and operating in a channel-cut'' mode. The second of the crystals can be bent for sagittal focusing of the beam. The second monochromator unit reflects 2 mrad of the radiation fan between the A2 and [ital B] branchesmore » of the beamline at a fixed angle of 2[Theta]=12[degree], and serves the sideways experimental station A1. Four interchangeable triangular crystals, mounted on a rotary holder, provide x rays of four different energies.« less