Lipidomic and in-gel analysis of maleic acid co-polymer nanodiscs reveals differences in composition of solubilized membranes

Barniol‐Xicota, Marta; Verhelst, Steven H. L.

doi:10.1038/s42003-021-01711-3

Cited by 24 publications

(35 citation statements)

References 49 publications

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“…The identification and quantification of lipids in nanodiscs, as shown here, is of particular interest for two reasons: First, the integration of different lipid classes and species into the nanodiscs during its assembly has to be carefully evaluated and verified to ensure that the desired membranes are used for subsequent experiments. Second, nanodisc polymers can be employed to solubilize natural lipid membranes; the identification and quantification of the lipid content in these membrane sections provides valuable insights into the composition of the natural membranes, including, for instance, the lipid environment of specific membrane proteins [24] . The analysis of the lipid content of nanodiscs was previously assessed by high‐resolution native MS [25] .…”

Section: Resultsmentioning

confidence: 99%

Thin‐Layer Chromatography and Coomassie Staining of Phospholipids for Fast and Simple Lipidomics Sample Preparation

et al. 2021

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Lipids play major roles in basic cellular functions. Their analysis, therefore, gained importance; however, it is complicated by the high complexity of natural lipidomes. To overcome this challenge, pre‐separation of the lipids by thin‐layer chromatography (TLC) and high‐resolution mass spectrometry (MS) are often employed. Here, we explore Coomassie staining for TLC‐separated phospholipids and provide an extraction protocol of stained lipids for subsequent MS analysis. We exemplify our approach by analyzing lipid mixtures varying in complexity and found that TLC pre‐separation increases the number of identified lipid species and lipid classes. In addition, we identify and quantify lipids from polymer nanodiscs. In summary, Coomassie staining of TLC‐separated lipids is well‐suited for phospholipids, is compatible with MS, does not require specialized equipment and can be performed independently of subsequent MS experiments. We envision many future applications of our workflow.

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Section: Resultsmentioning

confidence: 99%

Thin‐Layer Chromatography and Coomassie Staining of Phospholipids for Fast and Simple Lipidomics Sample Preparation

et al. 2021

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“…One study reported no differences in lipid content between SMALPs and the originating E. coli membrane [ 72 ], whereas subtle differences were observed between total membrane and SMA solubilised samples for Pichia pastoris expressing CD81 [ 22 ]. Lipidomic analysis of nanodiscs generated using a variety of polymers, including SMA and DIBMA, suggested that glycerolipids are strongly enriched and phospholipids de-enriched in polymer-solubilised samples compared with the originating E. coli and Jurkat membranes [ 75 ]. In model heterogenous phase-separated bilayers, SMA preferentially solubilised lipids in the fluid phase [ 76 ], suggesting that biological membrane microdomains with differing fluidity may be differentially soluble in SMA.…”

Section: Protein–lipid Interactionsmentioning

confidence: 99%

Biological insights from SMA-extracted proteins

Unger

Ronco-Campaña

Kitchen

et al. 2021

Biochemical Society Transactions

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In the twelve years since styrene maleic acid (SMA) was first used to extract and purify a membrane protein within a native lipid bilayer, this technological breakthrough has provided insight into the structural and functional details of protein–lipid interactions. Most recently, advances in cryo-EM have demonstrated that SMA-extracted membrane proteins are a rich-source of structural data. For example, it has been possible to resolve the details of annular lipids and protein–protein interactions within complexes, the nature of lipids within central cavities and binding pockets, regions involved in stabilising multimers, details of terminal residues that would otherwise remain unresolved and the identification of physiologically relevant states. Functionally, SMA extraction has allowed the analysis of membrane proteins that are unstable in detergents, the characterization of an ultrafast component in the kinetics of electron transfer that was not possible in detergent-solubilised samples and quantitative, real-time measurement of binding assays with low concentrations of purified protein. While the use of SMA comes with limitations such as its sensitivity to low pH and divalent cations, its major advantage is maintenance of a protein's lipid bilayer. This has enabled researchers to view and assay proteins in an environment close to their native ones, leading to new structural and mechanistic insights.

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“…These factors, collectively, influence polymer-polymer, polymer-lipid, protein-polymer interactions, which may compromise the yield, purity, activity of final purified target protein, and thus the downstream analyses. Not to mention that different polymers have shown to display differential preference for solubilization of specific types of lipids in biomembranes derived from prokaryotes and eukaryotes [ 58 ]. As demonstrated by multiple lines of research, the lipid composition of polymer-based nanodiscs is inevitably susceptible to change through inter-particle collision.…”

Section: Synthetic Polymers For Reconstitution Of Membrane Assembliesmentioning

confidence: 99%

Current Developments in Native Nanometric Discoidal Membrane Bilayer Formed by Amphipathic Polymers

2021

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Unlike cytosolic proteins, membrane proteins (MPs) are embedded within the plasma membrane and the lipid bilayer of intracellular organelles. MPs serve in various cellular processes and account for over 65% of the current drug targets. The development of membrane mimetic systems such as bicelles, short synthetic polymers or amphipols, and membrane scaffold proteins (MSP)-based nanodiscs has facilitated the accommodation of synthetic lipids to stabilize MPs, yet the preparation of these membrane mimetics remains detergent-dependent. Bio-inspired synthetic polymers present an invaluable tool for excision and liberation of superstructures of MPs and their surrounding annular lipid bilayer in the nanometric discoidal assemblies. In this article, we discuss the significance of self-assembling process in design of biomimetic systems, review development of multiple series of amphipathic polymers and the significance of these polymeric “belts” in biomedical research in particular in unraveling the structures, dynamics and functions of several high-value membrane protein targets.

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Lipidomic and in-gel analysis of maleic acid co-polymer nanodiscs reveals differences in composition of solubilized membranes

Cited by 24 publications

References 49 publications

Thin‐Layer Chromatography and Coomassie Staining of Phospholipids for Fast and Simple Lipidomics Sample Preparation

Thin‐Layer Chromatography and Coomassie Staining of Phospholipids for Fast and Simple Lipidomics Sample Preparation

Biological insights from SMA-extracted proteins

Current Developments in Native Nanometric Discoidal Membrane Bilayer Formed by Amphipathic Polymers

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