Engineering Nanodisc Scaffold Proteins for Native Mass Spectrometry

Reid, Deseree J.; Keener, James E.; Wheeler, Andrew P.; Zambrano, Dane Evan; Diesing, Jessica Marie; Reinhardt-Szyba, Maria; Makarov, Alexander; Marty, Michael T.

doi:10.1021/acs.analchem.7b03569

Cited by 52 publications

(99 citation statements)

References 29 publications

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“…Although the SoftMax function pushes the algorithm towards a single charge state for each data point, it allows for overlapping peaks where a data point can be legitimately assigned to more than one charge state. Finally, a spectrum of POPC nanodiscs with mixed membrane scaffold protein belts was investigated ( Figure 2g) [8]. Although well-resolved, this spectrum contains many overlapping peaks.…”

Section: Resultsmentioning

confidence: 99%

Eliminating Artifacts in Electrospray Deconvolution with a SoftMax Function

Marty

2019

J. Am. Soc. Mass Spectrom.

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UniDec provides a rapid and robust approach to deconvolving electrospray mass spectra into their corresponding mass and charge components. However, the UniDec algorithm can produce artifacts depending on the quality and complexity of the data. Here, a SoftMax function is applied to the charge state distribution of each data point, which pushes the algorithm towards assigning each data point to one primary charge state. As shown for several data sets of increasing complexity, the SoftMax function significantly reduces deconvolution artifacts, even for data with overlapping charge states.

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Section: Resultsmentioning

confidence: 99%

Eliminating Artifacts in Electrospray Deconvolution with a SoftMax Function

Marty

2019

J. Am. Soc. Mass Spectrom.

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“…110,111 A first study explored nanodiscs, bicelles and amphipols for native MS and showed that indeed the native oligomeric states of several integral membrane proteins were preserved in nanodiscs and bicelles while they differed from those obtained from detergent micelles or amphipols. 112 In particular, nanodiscs were therefore examined in detail; the complex and heterogeneous mass spectra further induced improvements in spectral deconvolution, [113][114][115][116] sample preparation [117][118][119] and method optimisation. 119,120 These improvements allowed, for instance, monitoring the insertion and oligomer assembly in the lipid bilayer.…”

Section: Unravelling the Architecture Of Membrane Protein Assembliementioning

confidence: 99%

Native mass spectrometry—A valuable tool in structural biology

Barth

Schmidt

2020

J Mass Spectrom

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Proteins and the complexes they form with their ligands are the players of cellular action. Their function is directly linked with their structure making the structural analysis of protein-ligand complexes essential. Classical techniques of structural biology include X-ray crystallography, nuclear magnetic resonance spectroscopy and recently distinguished cryo-electron microscopy. However, protein-ligand complexes are often dynamic and heterogeneous and consequently challenging for these techniques. Alternative approaches are therefore needed and gained importance during the last decades. One alternative is native mass spectrometry, which is the analysis of intact protein complexes in the gas phase. To achieve this, sample preparation and instrument conditions have to be optimised. Native mass spectrometry then reveals stoichiometry, protein interactions and topology of protein assemblies. Advanced techniques such as ion mobility and high-resolution mass spectrometry further add to the range of applications and deliver information on shape and microheterogeneity of the complexes. In this tutorial, we explain the basics of native mass spectrometry including sample requirements, instrument modifications and interpretation of native mass spectra. We further discuss the developments of native mass spectrometry and provide example spectra and applications.

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“…Such analyses could be leveraged to further understand how MPs interface with their surrounding lipid environment. Empty nanodiscs have also been studied by native MS allowing, for example, the precise lipid composition to be determined [117][118][119][120].…”

Section: Disc Technologies For Native Msmentioning

confidence: 99%

Mass spectrometry-enabled structural biology of membrane proteins

Calabrese

Radford

2018

Methods

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a b s t r a c tThe last $25 years has seen mass spectrometry (MS) emerge as an integral method in the structural biology toolkit. In particular, MS has enabled the structural characterization of proteins and protein assemblies that have been intractable by other methods, especially those that are large, heterogeneous or transient, providing experimental evidence for their structural organization in support of, and in advance of, high resolution methods. The most recent frontier conquered in the field of MS-based structural biology has been the application of established methods for studying water soluble proteins to the more challenging targets of integral membrane proteins. The power of MS in obtaining structural information has been enabled by advances in instrumentation and the development of hyphenated mass spectrometry-based methods, such as ion mobility spectrometry-MS, chemical crosslinking-MS and other chemical labelling/footprinting-MS methods. In this review we detail the insights garnered into the structural biology of membrane proteins by applying such techniques. Application and refinement of these methods has yielded unprecedented insights in many areas, including membrane protein conformation, dynamics, lipid/ligand binding, and conformational perturbations due to ligand binding, which can be challenging to study using other methods.

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Engineering Nanodisc Scaffold Proteins for Native Mass Spectrometry

Cited by 52 publications

References 29 publications

Eliminating Artifacts in Electrospray Deconvolution with a SoftMax Function

Eliminating Artifacts in Electrospray Deconvolution with a SoftMax Function

Native mass spectrometry—A valuable tool in structural biology

Mass spectrometry-enabled structural biology of membrane proteins

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