NMR as a tool to investigate the structure, dynamics and function of membrane proteins

Liang, Binyong; Tamm, Lukas K.

doi:10.1038/nsmb.3226

Cited by 103 publications

(87 citation statements)

References 84 publications

(105 reference statements)

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“…However, what distinguishes our structures from nanodiscs? Nanodiscs are defined as patches of a lipid bilayer surrounded by amphipathic helices of proteins or peptides [51]. Because V 4 D contains only 5 amino acid residues, we assume that it does not attain the critical length that renders helix formation possible.…”

Section: Peptide-induced Bicelle Formationmentioning

confidence: 99%

Peptide self-assembly into lamellar phases and the formation of lipid-peptide nanostructures

et al. 2017

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Lipids exhibit an extraordinary polymorphism in self-assembled mesophases, with lamellar phases as the most relevant biological representative. To mimic lipid lamellar phases with amphiphilic designer peptides, seven systematically varied short peptides were engineered. Indeed, four peptide candidates (V 4 D, V 4 WD, V 4 WD 2 , I 4 WD 2 ) readily self-assembled into lamellae in aqueous solution. Small-angle X-ray scattering (SAXS) patterns revealed ordered lamellar structures with a repeat distance of ~ 4-5 nm. Transmission electron microscopy (TEM) images confirmed the presence of stacked sheets. Two derivatives (V 3 D and V 4 D 2 ) remained as loose aggregates dispersed in solution; one peptide (L 4 WD 2 ) formed twisted tapes with internal lamellae and an antiparallel β-type monomer alignment. To understand the interaction of peptides with lipids, they were mixed with phosphatidylcholines. Low peptide concentrations (1.1 mM) induced the formation of a heterogeneous mixture of vesicular structures. Large multilamellar vesicles (MLV, d-spacing ~ 6.3 nm) coexisted with oligo-or unilamellar vesicles (~ 50 nm in diameter) and bicelle-like structures (~ 45 nm length, ~ 18 nm width). High peptide concentrations (11 mM) led to unilamellar vesicles (ULV, diameter ~ 260-280 nm) with a homogeneous mixing of lipids and peptides. SAXS revealed the temperature-dependent fine structure of these ULVs. At 25 °C the bilayer is in a fully interdigitated state (headgroup-to-headgroup distance d HH ~ 2.9 nm), whereas at 50 °C this interdigitation opens up (d HH ~ 3.6 nm). Our results highlight the versatility of self-assembled peptide superstructures. Subtle changes in the amino acid composition are key design elements in creating peptide-or lipidpeptide nanostructures with richness in morphology similar to that of naturally occurring lipids.

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Section: Peptide-induced Bicelle Formationmentioning

confidence: 99%

Peptide self-assembly into lamellar phases and the formation of lipid-peptide nanostructures

et al. 2017

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“…Here again, however, data from other methods, such as single molecule Förster Resonance Energy Transfer (FRET) [244,245], small angle X-ray scattering (SAXS) [246,247], and double electron-electron resonance (DEER) electron paramagnetic resonance (EPR) [248], can provide independent information that complements MS data so that the two approaches can be used in synergy (so-called 'integrated structural biology' [249]) to create molecular models. This issue is not unique to MS footprinting methods, with structural techniques such as NMR and cryo-EM also potentially being resolution limited by dynamic averaging [13][14][15]250,251]. Powerful NMR methods, such as relaxation dispersion, however, can provide structural, kinetic and thermodynamic information about rare species in dynamic equilibrium, in favorable cases even for large (MDa) protein complexes [250,251].…”

Section: Discussionmentioning

confidence: 99%

“…This issue is not unique to MS footprinting methods, with structural techniques such as NMR and cryo-EM also potentially being resolution limited by dynamic averaging [13][14][15]250,251]. Powerful NMR methods, such as relaxation dispersion, however, can provide structural, kinetic and thermodynamic information about rare species in dynamic equilibrium, in favorable cases even for large (MDa) protein complexes [250,251]. In the same vein, particle classification methods in cryo-EM can be used to tease out different protein structures within a dynamic ensemble provided that each conformer is significantly represented so that medium to high resolution data can be obtained [13][14][15].…”

Section: Discussionmentioning

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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“…Solution NMR has been successfully used to study membrane proteins solubilized in different membrane mimicking systems, including organic solvent mixtures, amphipols, micelles, and bicelles [6, 7]. These media, though useful, present themselves with a number of caveats, including surface curvature artifacts, limited diversity of detergent or lipid molecules, heterogeneity of the sample preparation and a debilitating inability to study any interaction with their soluble binding partners.…”

Section: Introductionmentioning

confidence: 99%

Nanodiscs and solution NMR: preparation, application and challenges

Puthenveetil

Nguyen

Vinogradova

2017

Nanotechnology Reviews

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Nanodiscs provide an excellent system for the structure-function investigation of membrane proteins. Its direct advantage lies in presenting a water soluble form of an otherwise hydrophobic molecule, making it amenable to a plethora of solution techniques. Nuclear Magnetic Resonance is one such high resolution approach that looks at the structure and dynamics of a protein with atomic level precision. Recently, there has been a breakthrough in making nanodiscs more susceptible for structure determination by solution NMR, yet it still remains to become the preferred choice for a membrane mimetic. In this practical review, we provide a general discourse on nanodisc and its application to solution NMR. We also offer potential solutions to remediate the technical challenges associated with nanodisc preparation and the choice of proper experimental set-ups. Along with discussing several structural applications, we demonstrate an alternative use of nanodiscs for functional studies, where we investigated the phosphorylation of a cell surface receptor, Integrin. This is the first successful manifestation of observing activated receptor phosphorylation in nanodiscs through NMR. We additionally present an on-column method for nanodisc preparation with multiple strategies and discuss the potential use of alternative nanoscale phospholipid bilayer systems like SMA lipid discs and Saposin-A lipoprotein discs.

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NMR as a tool to investigate the structure, dynamics and function of membrane proteins

Cited by 103 publications

References 84 publications

Peptide self-assembly into lamellar phases and the formation of lipid-peptide nanostructures

Peptide self-assembly into lamellar phases and the formation of lipid-peptide nanostructures

Mass spectrometry-enabled structural biology of membrane proteins

Nanodiscs and solution NMR: preparation, application and challenges

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