Microfluidic sorting of protein nanocrystals by size for X-ray free-electron laser diffraction

Abdallah, Bahige G.; Zatsepin, Nadia A.; Roy-Chowdhury, Shatabdi; Coe, Jesse; Conrad, Chelsie E.; Dörner, Katerina; Sierra, Raymond G.; Stevenson, Hilary P.; Camacho-Alanis, Fernanda; Grant, Thomas D.; Nelson, Garrett; James, Daniel; Calero, Guillermo; Wachter, Rebekka M.; Spence, John C.  H.; Weierstall, Uwe; Fromme, Petra; Ros, Alexandra

doi:10.1063/1.4928688

Title: Microfluidic sorting of protein nanocrystals by size for X-ray free-electron laser diffraction

Journal Article · Wed Aug 19 00:00:00 EDT 2015 · Structural Dynamics

DOI:https://doi.org/10.1063/1.4928688· OSTI ID:1261128

Abdallah, Bahige G. ^[1]; Zatsepin, Nadia A. ^[2]; Roy-Chowdhury, Shatabdi ^[1]; Coe, Jesse ^[1]; Conrad, Chelsie E. ^[1]; Dörner, Katerina ^[3]; Sierra, Raymond G. ^[4]; Stevenson, Hilary P. ^[5]; Camacho-Alanis, Fernanda ^[6]; Grant, Thomas D. ^[7]; Nelson, Garrett ^[2]; James, Daniel ^[2]; Calero, Guillermo ^[5]; Wachter, Rebekka M. ^[6]; Spence, John C. H. ^[2]; Weierstall, Uwe ^[2]; Fromme, Petra ^[1]; Ros, Alexandra ^[1]

Arizona State Univ., Tempe, AZ (United States). Dept. of Chemistry and Biochemistry; Arizona State Univ., Tempe, AZ (United States). Biodesign Inst.
Arizona State Univ., Tempe, AZ (United States). Biodesign Inst.; Arizona State Univ., Tempe, AZ (United States). Dept. of Physics
Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany)
SLAC National Accelerator Lab., Menlo Park, CA (United States). Photon Ultrafast Laser Science and Engineering Inst. (PULSE)
Univ. of Pittsburgh School of Medicine, Pittsburgh, PA (United States). Dept. of Structural Biology
Arizona State Univ., Tempe, AZ (United States). Dept. of Chemistry and Biochemistry
Hauptman-Woodward Medical Research Inst. (HWI), Buffalo, NY (United States)

We report that the advent and application of the X-ray free-electron laser (XFEL) has uncovered the structures of proteins that could not previously be solved using traditional crystallography. While this new technology is powerful, optimization of the process is still needed to improve data quality and analysis efficiency. One area is sample heterogeneity, where variations in crystal size (among other factors) lead to the requirement of large data sets (and thus 10–100 mg of protein) for determining accurate structure factors. To decrease sample dispersity, we developed a high-throughput microfluidic sorter operating on the principle of dielectrophoresis, whereby polydisperse particles can be transported into various fluid streams for size fractionation. Using this microsorter, we isolated several milliliters of photosystem I nanocrystal fractions ranging from 200 to 600 nm in size as characterized by dynamic light scattering, nanoparticle tracking, and electron microscopy. Sorted nanocrystals were delivered in a liquid jet via the gas dynamic virtual nozzle into the path of the XFEL at the Linac Coherent Light Source. We obtained diffraction to ~4 Å resolution, indicating that the small crystals were not damaged by the sorting process. We also observed the shape transforms of photosystem I nanocrystals, demonstrating that our device can optimize data collection for the shape transform-based phasing method. Using simulations, we show that narrow crystal size distributions can significantly improve merged data quality in serial crystallography. From this proof-of-concept work, we expect that the automated size-sorting of protein crystals will become an important step for sample production by reducing the amount of protein needed for a high quality final structure and the development of novel phasing methods that exploit inter-Bragg reflection intensities or use variations in beam intensity for radiation damage-induced phasing. Ultimately, this method will also permit an analysis of the dependence of crystal quality on crystal size.

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Research Organization:: SLAC National Accelerator Laboratory (SLAC), Menlo Park, CA (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Basic Energy Sciences (BES)

Grant/Contract Number:: R01GM095583; R01GM112686; R01GM097463; AC02-76SF00515; 1231306

OSTI ID:: 1261128

Alternate ID(s):: OSTI ID: 1421055

Journal Information:: Structural Dynamics, Vol. 2, Issue 4; ISSN 2329-7778

Publisher:: American Crystallographic Association/AIPCopyright Statement

Country of Publication:: United States

Language:: English

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Cited by: 22 works

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Microfluidics: From crystallization to serial time-resolved crystallography Sui, Shuo; Perry, Sarah L. Structural Dynamics, Vol. 4, Issue 3 https://doi.org/10.1063/1.4979640	journal	May 2017

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Title: Microfluidic sorting of protein nanocrystals by size for X-ray free-electron laser diffraction

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