Reversible stacking of lipid nanodiscs for structural studies of clotting factors

Nanodiscs (ND) are discoidal phospholipid bilayers stabilized by a pair of membrane-scaffolding proteins (MSP). The macromolecular composition and size of ND are ideal for structural and functional studies of membrane and membrane-associated proteins. In this work, we investigate the assembly of ND...

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Bibliographic Details
Published in:Nanotechnology reviews (Berlin) 2017-02, Vol.6 (1), p.139-148
Main Authors: Grushin, Kirill, White, Mark Andrew, Stoilova-McPhie, Svetla
Format: Article
Language:English
Subjects:
Blood coagulation
Clotting
Composition
cryo-electron microscopy
Electron microscopy
Ethylenediaminetetraacetic acids
Galactosylceramide
Lipid composition
lipid nanodiscs
Lipids
Macromolecules
Membranes
Phosphatidylserine
Phospholipids
Proteins
Scaffolding
Stacking
stacking of nanodiscs
Stacks
Structure-function relationships
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Summary:Nanodiscs (ND) are discoidal phospholipid bilayers stabilized by a pair of membrane-scaffolding proteins (MSP). The macromolecular composition and size of ND are ideal for structural and functional studies of membrane and membrane-associated proteins. In this work, we investigate the assembly of ND from a galactosylceramide and dioleoyl phosphatidylserine (PS) lipid mixture with two different MSP and at four MSP-to-lipid ratios. This lipid composition has been optimized for structural and biophysical studies of membrane-bound blood clotting factors that require Ca ions for function. We have demonstrated that CaCl induces reversible stacking of the ND that depends on the ND size and Ca concentrations. Our biophysical and electron microscopy (EM) studies show a predominant ND population of ~12 nm in diameter for both the ND assembled from MSP1D1 to lipids ratio of 1:40 and from MSP1E3D1-to-lipids ratio of 1:80. Approximately half of the ND population assembled at MSP1E3D1-to-lipids ratio of 1:150 has a diameter of ~16 nm. These larger ND form ordered stacks at 5-m Ca concentrations, as shown by cryo-EM. The number and length of the ND stacks increase with the increasing in Ca concentration. Adding millimolar concentrations of EDTA reverses the stacking of the ND.
ISSN:2191-9089
2191-9097
DOI:10.1515/ntrev-2016-0073