title: Serial crystallography captures enzyme catalysis in copper nitrite reductase at atomic resolution from one crystal.
abstract: Relating individual protein crystal structures to an enzyme mechanism remains a major and challenging goal for structural biology. Serial crystallography using multiple crystals has recently been reported in both synchrotron-radiation and X-ray free-electron laser experiments. In this work, serial crystallography was used to obtain multiple structures serially from one crystal (MSOX) to study in crystallo enzyme catalysis. Rapid, shutterless X-ray detector technology on a synchrotron MX beamline was exploited to perform low-dose serial crystallography on a single copper nitrite reductase crystal, which survived long enough for 45 consecutive 100 K X-ray structures to be collected at 1.07-1.62 Å resolution, all sampled from the same crystal volume. This serial crystallography approach revealed the gradual conversion of the substrate bound at the catalytic type 2 Cu centre from nitrite to nitric oxide, following reduction of the type 1 Cu electron-transfer centre by X-ray-generated solvated electrons. Significant, well defined structural rearrangements in the active site are evident in the series as the enzyme moves through its catalytic cycle, namely nitrite reduction, which is a vital step in the global denitrification process. It is proposed that such a serial crystallography approach is widely applicable for studying any redox or electron-driven enzyme reactions from a single protein crystal. It can provide a 'catalytic reaction movie' highlighting the structural changes that occur during enzyme catalysis. The anticipated developments in the automation of data analysis and modelling are likely to allow seamless and near-real-time analysis of such data on-site at some of the powerful synchrotron crystallographic beamlines.
authors.affiliation: School of Biological Sciences, University of Essex , Wivenhoe Park, Colchester CO4 3SQ, England.
authors.forename: Sam
authors.initials: S
authors.lastname: Horrell
authors.affiliation: Molecular Biophysics Group, Institute of Integrative Biology, University of Liverpool , Life Sciences Building, Liverpool L69 7ZB, England.
authors.forename: Svetlana V
authors.initials: SV
authors.lastname: Antonyuk
authors.affiliation: Molecular Biophysics Group, Institute of Integrative Biology, University of Liverpool , Life Sciences Building, Liverpool L69 7ZB, England.
authors.forename: Robert R
authors.initials: RR
authors.lastname: Eady
authors.affiliation: Molecular Biophysics Group, Institute of Integrative Biology, University of Liverpool , Life Sciences Building, Liverpool L69 7ZB, England.
authors.forename: S Samar
authors.initials: SS
authors.lastname: Hasnain
authors.affiliation: School of Biological Sciences, University of Essex , Wivenhoe Park, Colchester CO4 3SQ, England.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Hough
authors.affiliation: School of Biological Sciences, University of Essex , Wivenhoe Park, Colchester CO4 3SQ, England.
authors.forename: Richard W
authors.initials: RW
authors.lastname: Strange
keywords: MSOX; XFEL; catalysis; copper nitrite reductase; denitrification; enzyme mechanism; radiation damage; radiolysis; serial crystallography; synchrotron radiation