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

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

    Research output: Contribution to journalArticle

    7 Citations (Scopus)

    Abstract

    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. This method will also permit an analysis of the dependence of crystal quality on crystal size.

    Original languageEnglish (US)
    Article number041719
    JournalStructural Dynamics
    Volume2
    Issue number4
    DOIs
    StatePublished - Jul 1 2015

    Fingerprint

    X ray lasers
    Free electron lasers
    classifying
    Sorting
    Microfluidics
    free electron lasers
    Nanocrystals
    nanocrystals
    Diffraction
    proteins
    Proteins
    Crystals
    diffraction
    crystals
    Photosystem I Protein Complex
    x rays
    Crystallography
    crystallography
    Gas dynamics
    gas dynamics

    ASJC Scopus subject areas

    • Condensed Matter Physics
    • Instrumentation
    • Radiation
    • Spectroscopy

    Cite this

    Microfluidic sorting of protein nanocrystals by size for X-ray free-electron laser diffraction. / Abdallah, Bahige G.; Zatsepin, Nadia; 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; Spence, John; Weierstall, Uwe; Fromme, Petra; Ros, Alexandra.

    In: Structural Dynamics, Vol. 2, No. 4, 041719, 01.07.2015.

    Research output: Contribution to journalArticle

    Abdallah, BG, Zatsepin, N, Roy-Chowdhury, S, Coe, J, Conrad, CE, Dörner, K, Sierra, RG, Stevenson, HP, Camacho-Alanis, F, Grant, TD, Nelson, G, James, D, Calero, G, Wachter, R, Spence, J, Weierstall, U, Fromme, P & Ros, A 2015, 'Microfluidic sorting of protein nanocrystals by size for X-ray free-electron laser diffraction' Structural Dynamics, vol. 2, no. 4, 041719. https://doi.org/10.1063/1.4928688
    Abdallah, Bahige G. ; Zatsepin, Nadia ; 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 ; Spence, John ; Weierstall, Uwe ; Fromme, Petra ; Ros, Alexandra. / Microfluidic sorting of protein nanocrystals by size for X-ray free-electron laser diffraction. In: Structural Dynamics. 2015 ; Vol. 2, No. 4.
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