title: Development of a microsecond X-ray protein footprinting facility at the Advanced Light Source.
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.
authors: Berkeley Center for Structural Biology, Physical Biosciences Division, Lawrence Berkeley National Laboratory, 1 Cyclotron Road, Berkeley, CA 94720, USA.
authors: Sayan
authors: S
authors: Gupta
authors: Experimental Systems, Advanced Light Source Division, Lawrence Berkeley National Laboratory, 1 Cyclotron Road, Berkeley, CA 94720, USA.
authors: Richard
authors: R
authors: Celestre
authors: Joint BioEnergy Institute, Physical Biosciences Division, Lawrence Berkeley National Laboratory, 1 Cyclotron Road, Berkeley, CA 94720, USA.
authors: Christopher J
authors: CJ
authors: Petzold
authors: Center for Synchrotron Biosciences, Center for Proteomics and Bioinformatics, School of Medicine, Case Western Reserve University, Cleveland, OH 44106, USA.
authors: Mark R
authors: MR
authors: Chance
authors: Berkeley Center for Structural Biology, Physical Biosciences Division, Lawrence Berkeley National Laboratory, 1 Cyclotron Road, Berkeley, CA 94720, USA.
authors: Corie
authors: C
authors: Ralston
mesh_terms: D001665:Binding Sites; D018360:Crystallography, X-Ray / Q000295:instrumentation*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D011485:Protein Binding; D011487:Protein Conformation; D023302:Protein Footprinting / Q000295:instrumentation*; D011506:Proteins / Q000737:chemistry* / Q000648:ultrastructure*; D017356:Synchrotrons / Q000295:instrumentation*
chemical_list: D011506:Proteins
keywords: mass spectrometry; microsecond irradiation; protein structure; radiolytic labeling