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Article

Crystallization of ApoA1 and ApoE4 Nanolipoprotein Particles and Initial XFEL-Based Structural Studies

1
Biosciences and Biotechnology Division, Lawrence Livermore National Laboratory, Livermore, CA 94550, USA
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Department of Chemical Engineering, University of California at Davis, Davis, CA 95616, USA
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Department of Structural Biology, Jacobs School of Medicine and Biomedical Sciences, SUNY University at Buffalo, Buffalo, NY 14214, USA
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Hauptman-Woodward Medical Research Institute, Buffalo, NY 14203, USA
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Department of Radiation Oncology, School of Medicine, University of California at Davis, Sacramento, CA 95817, USA
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Center for Free-Electron Laser Science, Deutsches Elektronen-Synchrotron, 22607 Hamburg, Germany
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Linac Coherent Light Source, SLAC National Accelerator Laboratory, Menlo Park, CA 94025, USA
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Department of Biochemistry and Molecular Medicine, School of Medicine, University of California at Davis, Sacramento, CA 95817, USA
*
Authors to whom correspondence should be addressed.
Crystals 2020, 10(10), 886; https://doi.org/10.3390/cryst10100886
Received: 2 September 2020 / Revised: 23 September 2020 / Accepted: 23 September 2020 / Published: 1 October 2020
(This article belongs to the Special Issue Macromolecular Serial Crystallography (Volume II))
Nanolipoprotein particles (NLPs), also called “nanodiscs”, are discoidal particles with a patch of lipid bilayer corralled by apolipoproteins. NLPs have long been of interest due to both their utility as membrane-model systems into which membrane proteins can be inserted and solubilized and their physiological role in lipid and cholesterol transport via high-density lipoprotein (HDL) and low-density lipoprotein (LDL) maturation, which are important for human health. Serial femtosecond crystallography (SFX) at X-ray free electron lasers (XFELs) is a powerful approach for structural biology of membrane proteins, which are traditionally difficult to crystallize as large single crystals capable of producing high-quality diffraction suitable for structure determination. To facilitate understanding of the specific role of two apolipoprotein/lipid complexes, ApoA1 and ApoE4, in lipid binding and HDL/LDL particle maturation dynamics, and to develop new SFX methods involving NLP membrane protein encapsulation, we have prepared and crystallized homogeneous populations of ApoA1 and ApoE4 NLPs. Crystallization of empty NLPs yields semi-ordered objects that appear crystalline and give highly anisotropic and diffuse X-ray diffraction, similar to fiber diffraction. Several unit cell parameters were approximately determined for both NLPs from these measurements. Thus, low-background, sample conservative methods of delivery are critical. Here we implemented a fixed target sample delivery scheme utilizing the Roadrunner fast-scanning system and ultra-thin polymer/graphene support films, providing a low-volume, low-background approach to membrane protein SFX. This study represents initial steps in obtaining structural information for ApoA1 and ApoE4 NLPs and developing this system as a supporting scaffold for future structural studies of membrane proteins crystalized in a native lipid environment. View Full-Text
Keywords: lipoprotein; nanodisc; serial femtosecond crystallography; XFELs; fixed target delivery lipoprotein; nanodisc; serial femtosecond crystallography; XFELs; fixed target delivery
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MDPI and ACS Style

Shelby, M.L.; Gilbile, D.; Grant, T.D.; Bauer, W.J.; Segelke, B.; He, W.; Evans, A.C.; Crespo, N.; Fischer, P.; Pakendorf, T.; Hennicke, V.; Hunter, M.S.; Batyuk, A.; Barthelmess, M.; Meents, A.; Kuhl, T.L.; Frank, M.; Coleman, M.A. Crystallization of ApoA1 and ApoE4 Nanolipoprotein Particles and Initial XFEL-Based Structural Studies. Crystals 2020, 10, 886. https://doi.org/10.3390/cryst10100886

AMA Style

Shelby ML, Gilbile D, Grant TD, Bauer WJ, Segelke B, He W, Evans AC, Crespo N, Fischer P, Pakendorf T, Hennicke V, Hunter MS, Batyuk A, Barthelmess M, Meents A, Kuhl TL, Frank M, Coleman MA. Crystallization of ApoA1 and ApoE4 Nanolipoprotein Particles and Initial XFEL-Based Structural Studies. Crystals. 2020; 10(10):886. https://doi.org/10.3390/cryst10100886

Chicago/Turabian Style

Shelby, Megan L., Deepshika Gilbile, Thomas D. Grant, William J. Bauer, Brent Segelke, Wei He, Angela C. Evans, Natalia Crespo, Pontus Fischer, Tim Pakendorf, Vincent Hennicke, Mark S. Hunter, Alex Batyuk, Miriam Barthelmess, Alke Meents, Tonya L. Kuhl, Matthias Frank, and Matthew A. Coleman 2020. "Crystallization of ApoA1 and ApoE4 Nanolipoprotein Particles and Initial XFEL-Based Structural Studies" Crystals 10, no. 10: 886. https://doi.org/10.3390/cryst10100886

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