Rapid preparation of nanodiscs for biophysical studies.

Autor: Julien JA; Department of Chemistry, Lehigh University, 6 E. Packer Ave. Bethlehem, Pennsylvania, 18015, USA., Fernandez MG; Department of Chemistry, Lehigh University, 6 E. Packer Ave. Bethlehem, Pennsylvania, 18015, USA., Brandmier KM; Department of Chemistry, Lehigh University, 6 E. Packer Ave. Bethlehem, Pennsylvania, 18015, USA., Del Mundo JT; Department of Chemical Engineering, The Pennsylvania State University, 121 Chemical and Biomedical Engineering Building, University Park, PA, 16802, USA., Bator CM; Huck Institutes of Life Sciences, Cryo-EM Facility, The Pennsylvania State University, University Park, PA, 16802, USA., Loftus LA; Department of Chemistry, Lehigh University, 6 E. Packer Ave. Bethlehem, Pennsylvania, 18015, USA., Gomez EW; Department of Chemical Engineering, The Pennsylvania State University, 121 Chemical and Biomedical Engineering Building, University Park, PA, 16802, USA., Gomez ED; Department of Chemical Engineering, The Pennsylvania State University, 121 Chemical and Biomedical Engineering Building, University Park, PA, 16802, USA; Department of Materials Science and Engineering, The Pennsylvania State University, 404 Steidle Building, University Park, PA, 16802, USA., Glover KJ; Department of Chemistry, Lehigh University, 6 E. Packer Ave. Bethlehem, Pennsylvania, 18015, USA. Electronic address: kjg206@lehigh.edu.
Jazyk: angličtina
Zdroj: Archives of biochemistry and biophysics [Arch Biochem Biophys] 2021 Nov 15; Vol. 712, pp. 109051. Date of Electronic Publication: 2021 Oct 02.
DOI: 10.1016/j.abb.2021.109051
Abstrakt: Nanodiscs, which are disc-shaped entities that contain a central lipid bilayer encased by an annulus of amphipathic helices, have emerged as a leading native-like membrane mimic. The current approach for the formation of nanodiscs involves the creation of a mixed-micellar solution containing membrane scaffold protein, lipid, and detergent followed by a time consuming process (3-12 h) of dialysis and/or incubation with sorptive beads to remove the detergent molecules from the sample. In contrast, the methodology described herein provides a facile and rapid procedure for the preparation of nanodiscs in a matter of minutes (<15 min) using Sephadex® G-25 resin to remove the detergent from the sample. A panoply of biophysical techniques including analytical ultracentrifugation, dynamic light scattering, gel filtration chromatography, circular dichroism spectroscopy, and cryogenic electron microscopy were employed to unequivocally confirm that aggregates formed by this method are indeed nanodiscs. We believe that this method will be attractive for time-sensitive and high-throughput experiments.
(Copyright © 2021 Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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