Ultrasonic acoustic levitation for fast frame rate X-ray protein crystallography at room temperature

Increasing the data acquisition rate of X-ray diffraction images for macromolecular crystals at room temperature at synchrotrons has the potential to significantly accelerate both structural analysis of biomolecules and structure-based drug developments. Using lysozyme model crystals, we demonstrated the rapid acquisition of X-ray diffraction datasets by combining a high frame rate pixel array detector with ultrasonic acoustic levitation of protein crystals in liquid droplets. The rapid spinning of the crystal within a levitating droplet ensured an efficient sampling of the reciprocal space. The datasets were processed with a program suite developed for serial femtosecond crystallography (SFX). The structure, which was solved by molecular replacement, was found to be identical to the structure obtained by the conventional oscillation method for up to a 1.8-Å resolution limit. In particular, the absence of protein crystal damage resulting from the acoustic levitation was carefully established. These results represent a key step towards a fully automated sample handling and measurement pipeline, which has promising prospects for a high acquisition rate and high sample efficiency for room temperature X-ray crystallography.

[1]  Anton Barty,et al.  Room-temperature macromolecular serial crystallography using synchrotron radiation , 2014, IUCrJ.

[2]  Thomas Laurell,et al.  Screening of nucleation conditions using levitated drops for protein crystallization. , 2003, Analytical chemistry.

[3]  Staffan Nilsson,et al.  Airborne chemistry: acoustic levitation in chemical analysis , 2004, Analytical and bioanalytical chemistry.

[4]  Ezequiel Panepucci,et al.  Room-temperature serial crystallography at synchrotron X-ray sources using slowly flowing free-standing high-viscosity microstreams. , 2015, Acta crystallographica. Section D, Biological crystallography.

[5]  W. J. Xie,et al.  Acoustic method for levitation of small living animals , 2006 .

[6]  Roberto Dinapoli,et al.  PILATUS: A single photon counting pixel detector for X-ray applications , 2009 .

[7]  Edward Hæggström,et al.  Effects of acoustic levitation on the development of zebrafish, Danio rerio, embryos , 2015, Scientific Reports.

[8]  Geoffrey Lee,et al.  Surface temperature of acoustically levitated water microdroplets measured using infra-red thermography , 2008 .

[9]  S. Sadhal,et al.  Internal circulation in a drop in an acoustic field. , 1999, Journal of the Acoustical Society of America.

[10]  Acoustically mounted microcrystals yield high-resolution X-ray structures. , 2011, Biochemistry.

[11]  W. Xie,et al.  Containerless solidification of acoustically levitated Ni–Sn eutectic alloy , 2012 .

[12]  Jork Leiterer,et al.  Agglomeration of proteins in acoustically levitated droplets , 2008, Analytical and bioanalytical chemistry.

[13]  G. Brenn,et al.  Evaporation of acoustically levitated droplets , 1999, Journal of Fluid Mechanics.

[14]  T. Tomizaki,et al.  SLS Crystallization Platform at Beamline X06DA—A Fully Automated Pipeline Enabling in Situ X-ray Diffraction Screening , 2011 .

[15]  Anton Barty,et al.  CrystFEL: a software suite for snapshot serial crystallography , 2012 .

[16]  D. McNaughton,et al.  Chemical analysis of acoustically levitated drops by Raman spectroscopy , 2009, Analytical and bioanalytical chemistry.

[17]  Louis Vessot King,et al.  On the Acoustic Radiation Pressure on Spheres , 1934 .

[18]  W. Xie,et al.  Levitation of iridium and liquid mercury by ultrasound. , 2002, Physical review letters.

[19]  R. Thorne,et al.  Global radiation damage at 300 and 260 K with dose rates approaching 1 MGy s⁻¹. , 2012, Acta crystallographica. Section D, Biological crystallography.

[20]  J. L. Robey,et al.  Experimental study of streaming flows associated with ultrasonic levitators , 1994 .

[21]  Alexander Scheeline,et al.  Potential of levitated drops to serve as microreactors for biophysical measurements. , 2012, Biophysical chemistry.

[22]  Gwyndaf Evans,et al.  Outrunning free radicals in room-temperature macromolecular crystallography , 2012, Acta crystallographica. Section D, Biological crystallography.

[23]  Nathaniel Echols,et al.  Accessing protein conformational ensembles using room-temperature X-ray crystallography , 2011, Proceedings of the National Academy of Sciences.

[24]  Jesse B. Hopkins,et al.  Figures and figure supplements Mapping the conformational landscape of a dynamic enzyme by multitemperature and XFEL crystallography , 2016 .

[25]  Gwyndaf Evans,et al.  In situ macromolecular crystallography using microbeams , 2012, Acta crystallographica. Section D, Biological crystallography.

[26]  Randy J. Read,et al.  Overview of the CCP4 suite and current developments , 2011, Acta crystallographica. Section D, Biological crystallography.

[27]  K. Hasegawa,et al.  Internal and External Flow Structure and Mass Transport Phenomena of an Acoustically Levitated Droplet , 2012 .

[28]  M. Selmer,et al.  Structure of the 70S Ribosome Complexed with mRNA and tRNA , 2006, Science.

[29]  S. McNicholas,et al.  Presenting your structures: the CCP4mg molecular-graphics software , 2011, Acta crystallographica. Section D, Biological crystallography.

[30]  B. Schmitt,et al.  EIGER: Next generation single photon counting detector for X-ray applications , 2011 .

[31]  Ezequiel Panepucci,et al.  In meso in situ serial X-ray crystallography of soluble and membrane proteins , 2015, Acta crystallographica. Section D, Biological crystallography.

[32]  R. Sweet,et al.  Hitting the target: fragment screening with acoustic in situ co-crystallization of proteins plus fragment libraries on pin-mounted data-collection micromeshes , 2014, Acta crystallographica. Section D, Biological crystallography.

[33]  Dimos Poulikakos,et al.  Acoustophoretic contactless transport and handling of matter in air , 2013, Proceedings of the National Academy of Sciences.

[34]  U. Panne,et al.  Structure analysis using acoustically levitated droplets , 2008, Analytical and bioanalytical chemistry.

[35]  W. Xie,et al.  Observation of ice nucleation in acoustically levitated water drops , 2005 .

[36]  A. Vagin,et al.  MOLREP: an Automated Program for Molecular Replacement , 1997 .

[37]  K. Aoki,et al.  Study on Internal Flow and Surface Deformation of Large Droplet Levitated by Ultrasonic Wave , 2006, Annals of the New York Academy of Sciences.

[38]  Zakiah N. Pierre,et al.  Sample Handling and Chemical Kinetics in an Acoustically Levitated Drop Microreactor , 2009, Analytical chemistry.

[39]  G. Murshudov,et al.  Refinement of macromolecular structures by the maximum-likelihood method. , 1997, Acta crystallographica. Section D, Biological crystallography.

[40]  Florent Cipriani,et al.  CrystalDirect: a new method for automated crystal harvesting based on laser-induced photoablation of thin films. , 2012, Acta crystallographica. Section D, Biological crystallography.

[41]  Jun Rekimoto,et al.  Three-Dimensional Mid-Air Acoustic Manipulation by Ultrasonic Phased Arrays , 2013, PloS one.

[42]  W. Burmeister,et al.  Structural changes in a cryo-cooled protein crystal owing to radiation damage. , 2000, Acta crystallographica. Section D, Biological crystallography.

[43]  Chris J. Benmore,et al.  Acoustic levitation: recent developments and emerging opportunities in biomaterials research , 2011, European Biophysics Journal.

[44]  Alexei S Soares,et al.  Acoustic methods for high-throughput protein crystal mounting at next-generation macromolecular crystallographic beamlines. , 2013, Journal of synchrotron radiation.

[45]  Sébastien Boutet,et al.  Protein crystal structure obtained at 2.9 Å resolution from injecting bacterial cells into an X-ray free-electron laser beam , 2014, Proceedings of the National Academy of Sciences.