Detergent-free Lipodisq Nanoparticles Facilitate High-Resolution Mass Spectrometry of Folded Integral Membrane Proteins

Hoi, Kin Kuan; Juarez, Juan F. Bada; Judge, Peter J.; Yen, Hsin‐Yung; Wu, Di; Vinals, Javier; Taylor, Garrick F.; Watts, Anthony; Robinson, Carol V.

doi:10.1021/acs.nanolett.0c04911

Cited by 30 publications

(24 citation statements)

References 79 publications

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“…For example, native mass spectrometry of two different bacteriorhodopsins extracted using SMA showed the presence of native ether-linked lipids. For one of the proteins this was also observed when the detergent octyl glucoside was used, but for the other this lipid adduct was lost with detergent [ 67 ]. Thus, it seems more likely that lipid-interactions will be maintained when using SMA than conventional detergents.…”

Section: Protein–lipid Interactionsmentioning

confidence: 99%

“…This has the potential for future structural and biophysical analysis to gain further insight to the mechanism of co-translational membrane protein folding and membrane insertion. Recently, native-mass spectrometry of SMALP-encapsulated proteins has also been demonstrated, enabling investigation of post-translational modifications, association with specific lipids and protein maturation [ 67 ]. Fluorescence correlation spectroscopy of SMALP purified proteins has been utilised to develop a new method for measuring ligand binding to membrane proteins, including both an ABC transporter [ 49 ], and a GPCR [ 68 ].…”

Section: Functional Insightsmentioning

confidence: 99%

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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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Section: Protein–lipid Interactionsmentioning

confidence: 99%

Section: Functional Insightsmentioning

confidence: 99%

Biological insights from SMA-extracted proteins

Unger

Ronco-Campaña

Kitchen

et al. 2021

Biochemical Society Transactions

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“…The endogenous lipid interactions of archeorhodopsin and bacteriorhodopsin, which are lost with detergent, are maintained in SMALPs. Native mass spectrometry revealed that these receptors complexed with diphytanyl glycerol lipid ligands (S-DGD and 2DP) in native membranes isolated using SMA(3:1) copolymer and DMPC [ 56 ]. This study compared n-octyl-β-d-glucoside detergent preparations and membrane scaffold protein (MSP)-based nanodiscs and showed that SMALPs are uniquely suited to solubilizing and stabilizing the correctly folded form of the protein while retaining bound biological lipids, which have long been known to preserve its stability and activity [ 70 ].…”

Section: Microbial Rhodopsins In Native Nanodiscsmentioning

confidence: 99%

Structures and Dynamics of Native-State Transmembrane Protein Targets and Bound Lipids

et al. 2021

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Membrane proteins work within asymmetric bilayers of lipid molecules that are critical for their biological structures, dynamics and interactions. These properties are lost when detergents dislodge lipids, ligands and subunits, but are maintained in native nanodiscs formed using styrene maleic acid (SMA) and diisobutylene maleic acid (DIBMA) copolymers. These amphipathic polymers allow extraction of multicomponent complexes of post-translationally modified membrane-bound proteins directly from organ homogenates or membranes from diverse types of cells and organelles. Here, we review the structures and mechanisms of transmembrane targets and their interactions with lipids including phosphoinositides (PIs), as resolved using nanodisc systems and methods including cryo-electron microscopy (cryo-EM) and X-ray diffraction (XRD). We focus on therapeutic targets including several G protein-coupled receptors (GPCRs), as well as ion channels and transporters that are driving the development of next-generation native nanodiscs. The design of new synthetic polymers and complementary biophysical tools bodes well for the future of drug discovery and structural biology of native membrane:protein assemblies (memteins).

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“…Recently, the availability of MS instruments designed to bring bigger and more complex systems into the gas phase-coupled with dedicated software development for the analysis of IMPs embedded in nanodiscs (Unidec)-has facilitated nMS studies of proteins in nanodiscs [51,52] or SMALPs [53]. Such progress will lead to more research on lipid-mediated oligomerization in a detergent-free environment [54,55].…”

Section: Lipids As Regulators Of Protein Oligomerization and Complex Assemblymentioning

confidence: 99%

Assessing the Role of Lipids in the Molecular Mechanism of Membrane Proteins

Jodaitis

Oene

Martens

2021

IJMS

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Membrane proteins have evolved to work optimally within the complex environment of the biological membrane. Consequently, interactions with surrounding lipids are part of their molecular mechanism. Yet, the identification of lipid–protein interactions and the assessment of their molecular role is an experimental challenge. Recently, biophysical approaches have emerged that are compatible with the study of membrane proteins in an environment closer to the biological membrane. These novel approaches revealed specific mechanisms of regulation of membrane protein function. Lipids have been shown to play a role in oligomerization, conformational transitions or allosteric coupling. In this review, we summarize the recent biophysical approaches, or combination thereof, that allow to decipher the role of lipid–protein interactions in the mechanism of membrane proteins.

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Detergent-free Lipodisq Nanoparticles Facilitate High-Resolution Mass Spectrometry of Folded Integral Membrane Proteins

Cited by 30 publications

References 79 publications

Biological insights from SMA-extracted proteins

Biological insights from SMA-extracted proteins

Structures and Dynamics of Native-State Transmembrane Protein Targets and Bound Lipids

Assessing the Role of Lipids in the Molecular Mechanism of Membrane Proteins

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