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J Synchrotron Radiat. 2014 Jul;21(Pt 4):690-9. doi: 10.1107/S1600577514007000. Epub 2014 May 16.

Development of a microsecond X-ray protein footprinting facility at the Advanced Light Source.

Author information

1
Berkeley Center for Structural Biology, Physical Biosciences Division, Lawrence Berkeley National Laboratory, 1 Cyclotron Road, Berkeley, CA 94720, USA.
2
Experimental Systems, Advanced Light Source Division, Lawrence Berkeley National Laboratory, 1 Cyclotron Road, Berkeley, CA 94720, USA.
3
Joint BioEnergy Institute, Physical Biosciences Division, Lawrence Berkeley National Laboratory, 1 Cyclotron Road, Berkeley, CA 94720, USA.
4
Center for Synchrotron Biosciences, Center for Proteomics and Bioinformatics, School of Medicine, Case Western Reserve University, Cleveland, OH 44106, USA.

Abstract

X-ray footprinting (XF) is an important structural biology tool used to determine macromolecular conformations and dynamics of both nucleic acids and proteins in solution on a wide range of timescales. With the impending shut-down of the National Synchrotron Light Source, it is ever more important that this tool continues to be developed at other synchrotron facilities to accommodate XF users. Toward this end, a collaborative XF program has been initiated at the Advanced Light Source using the white-light bending-magnet beamlines 5.3.1 and 3.2.1. Accessibility of the microsecond time regime for protein footprinting is demonstrated at beamline 5.3.1 using the high flux density provided by a focusing mirror in combination with a micro-capillary flow cell. It is further reported that, by saturating samples with nitrous oxide, the radiolytic labeling efficiency is increased and the imprints of bound versus bulk water can be distinguished. These results both demonstrate the suitability of the Advanced Light Source as a second home for the XF experiment, and pave the way for obtaining high-quality structural data on complex protein samples and dynamics information on the microsecond timescale.

KEYWORDS:

mass spectrometry; microsecond irradiation; protein structure; radiolytic labeling

PMID:
24971962
PMCID:
PMC4073957
DOI:
10.1107/S1600577514007000
[Indexed for MEDLINE]
Free PMC Article

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