login Search Personalize Collections Help Observation of substrate diffusion and ligand binding in enzyme crystals using high-repetition-rate mix-and-inject serial crystallography Pandey, Suraj; Calvey, George; Katz, Andrea M.; Malla, Tek Narsingh; Koua, Faisal H. M.; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; 2021 Download Formats Add to Basket Files Action Filename Size Access Description License mf5055.pdf 2.3 MB Public - - Abstract Here, we illustrate what happens inside the catalytic cleft of an enzyme when substrate or ligand binds on single-millisecond timescales. The initial phase of the enzymatic cycle is observed with near-atomic resolution using the most advanced X-ray source currently available: the European XFEL (EuXFEL). The high repetition rate of the EuXFEL combined with our mix-and-inject technology enables the initial phase of ceftriaxone binding to the Mycobacterium tuberculosis β-lactamase to be followed using time-resolved crystallography in real time. It is shown how a diffusion coefficient in enzyme crystals can be derived directly from the X-ray data, enabling the determination of ligand and enzyme–ligand concentrations at any position in the crystal volume as a function of time. In addition, the structure of the irreversible inhibitor sulbactam bound to the enzyme at a 66 ms time delay after mixing is described. This demonstrates that the EuXFEL can be used as an important tool for biomedically relevant research. Details Title Observation of substrate diffusion and ligand binding in enzyme crystals using high-repetition-rate mix-and-inject serial crystallography Author Pandey, Suraj Calvey, George Katz, Andrea M. Malla, Tek Narsingh Koua, Faisal H. M. Martin-Garcia, Jose M. Poudyal, Ishwor Yang, Jay-How Vakili, Mohammad Yefanov, Oleksandr Zielinski, Kara A. Bajt, Sasa Awel, Salah Doerner, Katarina Frank, Matthias Gelisio, Luca Jernigan, Rebecca Kirkwood, Henry Kloos, Marco Chalappurath Payyan Koliyadu, Jayanath Mariani, Valerio Miller, Mitchell D. Mills, Grant Nelson, Garrett Olmos, Jose L. Sadri, Alireza Sato, Tokushi Tolstikova, Alexandra Xu, Weijun Ourmazd, Abbas Spence, John C. H. Schwander, Peter Barty, Anton Chapman, Henry N. Fromme, Petra Mancuso, Adrian Phillips, George N. Bean, Richard Pollack, Lois Schmidt, Marius Affiliation EuXFEL staff, EuXFEL user Author Group Instrument SPB/SFX Sample Environment Keywords substrate diffusion in crystals; antibiotic resistance; β-lactamases; enzyme kinetics; irreversible inhibition; mix-and-inject serial crystallography; serial femtosecond crystallography; European X-ray Free-Electron Laser; megahertz pulse-repetition rate; protein structure determination; drug discovery; ceftriaxone; sulbactam; X-ray crystallography; enzyme mechanisms Topic SPB/SFX experiment Scientific Area Structural biology and biocrystallography X-ray scattering, X-ray optics, and instrumentation techniques Imprint International Union of Crystallography, 2021 Date 2021 Publication Information IUCrJ, 8 (6), 878-895 (2021) Related external records DOI: 10.1107/S2052252521008125 Content Type Articles Language English External Website https://doi.org/10.1107/S2052252521008125 Record Appears in Groups > Scientific instruments and equipment > Sample Environment Topics > XFEL.EU experiments Instruments > SPB/SFX Beamlines > SASE1 Types > Articles Years > 2021 Preview Select file: mf5055 Downloads Statistics from to By Days By Months By Years Update No data available. 2020-11 2021-01 2021-05 2021-08 2021-11 0 2 4 6 8 10 12 14 16 Total Downloads Download Full History