Increasing complexity in small-angle X-ray and neutron scattering experiments: from biological membrane mimics to live cells

Semeraro, Enrico F.; Marx, Lisa; Frewein, Moritz P. K.; Pabst, Georg

doi:10.1039/c9sm02352f

Cited by 36 publications

(25 citation statements)

References 110 publications

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“…the common assumption in analytical SAXS models that the scattering function can be described as a product of the bilayer form factor and the structure factor of the vesicle shape (Kiselev et al, 2002(Kiselev et al, , 2006. We hope that this contribution will facilitate the analysis of scattering data, which is particularly relevant given the currently increasing complexity in smallangle scattering experiments (Semeraro et al, 2021). Beyond the specific effects considered here, we also want to develop the approach of simulating vesicle ensembles and subsequent 3D FFT in view of the analysis of more complex vesicles and vesicle shape transitions, as required for example in studies of synaptic vesicles.…”

Section: Introductionmentioning

confidence: 93%

The effect of polydispersity, shape fluctuations and curvature on small unilamellar vesicle small-angle X-ray scattering curves

et al. 2021

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Small unilamellar vesicles (20–100 nm diameter) are model systems for strongly curved lipid membranes, in particular for cell organelles. Routinely, small-angle X-ray scattering (SAXS) is employed to study their size and electron-density profile (EDP). Current SAXS analysis of small unilamellar vesicles (SUVs) often employs a factorization into the structure factor (vesicle shape) and the form factor (lipid bilayer electron-density profile) and invokes additional idealizations: (i) an effective polydispersity distribution of vesicle radii, (ii) a spherical vesicle shape and (iii) an approximate account of membrane asymmetry, a feature particularly relevant for strongly curved membranes. These idealizations do not account for thermal shape fluctuations and also break down for strong salt- or protein-induced deformations, as well as vesicle adhesion and fusion, which complicate the analysis of the lipid bilayer structure. Presented here are simulations of SAXS curves of SUVs with experimentally relevant size, shape and EDPs of the curved bilayer, inferred from coarse-grained simulations and elasticity considerations, to quantify the effects of size polydispersity, thermal fluctuations of the SUV shape and membrane asymmetry. It is observed that the factorization approximation of the scattering intensity holds even for small vesicle radii (∼30 nm). However, the simulations show that, for very small vesicles, a curvature-induced asymmetry arises in the EDP, with sizeable effects on the SAXS curve. It is also demonstrated that thermal fluctuations in shape and the size polydispersity have distinguishable signatures in the SAXS intensity. Polydispersity gives rise to low-q features, whereas thermal fluctuations predominantly affect the scattering at larger q, related to membrane bending rigidity. Finally, it is shown that simulation of fluctuating vesicle ensembles can be used for analysis of experimental SAXS curves.

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Section: Introductionmentioning

confidence: 93%

The effect of polydispersity, shape fluctuations and curvature on small unilamellar vesicle small-angle X-ray scattering curves

et al. 2021

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“…Single particle cryo-EM can subsequently be used to solve the structure of MPs in these membrane mimics [135,245]. Additionally, small angle X-ray and neutron scattering may provide complementary approaches to help understand the relationship between MPs in those models, while mass spectrometry could be used to look at MPs directly ejected from native membranes [246][247][248][249][250]. Cryo-electron tomography (cryo-ET) is perhaps one technology that will enable conformational changes on the atomic scale to be linked with dynamic and temporal changes observed across whole cells [251].…”

Section: Resolutionmentioning

confidence: 99%

Changes in Membrane Protein Structural Biology

Birch

Cheruvara

Gamage

et al. 2020

Biology

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Membrane proteins are essential components of many biochemical processes and are important pharmaceutical targets. Membrane protein structural biology provides the molecular rationale for these biochemical process as well as being a highly useful tool for drug discovery. Unfortunately, membrane protein structural biology is a difficult area of study due to low protein yields and high levels of instability especially when membrane proteins are removed from their native environments. Despite this instability, membrane protein structural biology has made great leaps over the last fifteen years. Today, the landscape is almost unrecognisable. The numbers of available atomic resolution structures have increased 10-fold though advances in crystallography and more recently by cryo-electron microscopy. These advances in structural biology were achieved through the efforts of many researchers around the world as well as initiatives such as the Membrane Protein Laboratory (MPL) at Diamond Light Source. The MPL has helped, provided access to and contributed to advances in protein production, sample preparation and data collection. Together, these advances have enabled higher resolution structures, from less material, at a greater rate, from a more diverse range of membrane protein targets. Despite this success, significant challenges remain. Here, we review the progress made and highlight current and future challenges that will be overcome.

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“…Obtaining quantitative insight on a live cell's ultrastructure using either SAXS or SANS is challenging, however. This is simply because both techniques provide a global average of the entire cellular content, making it very difficult to single out individual contributors (Semeraro et al, 2020). This can be addressed with extensive use of the contrast variation capabilities of SANS.…”

Section: Introductionmentioning

confidence: 99%

Evolution of the analytical scattering model of live Escherichia coli

Semeraro¹,

Marx²,

Mandl³

et al. 2021

J Appl Cryst

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A previously reported multi-scale model for (ultra-)small-angle X-ray (USAXS/SAXS) and (very) small-angle neutron scattering (VSANS/SANS) of live Escherichia coli was revised on the basis of compositional/metabolomic and ultrastructural constraints. The cellular body is modeled, as previously described, by an ellipsoid with multiple shells. However, scattering originating from flagella was replaced by a term accounting for the oligosaccharide cores of the lipopolysaccharide leaflet of the outer membrane including its cross-term with the cellular body. This was mainly motivated by (U)SAXS experiments showing indistinguishable scattering for bacteria in the presence and absence of flagella or fimbrae. The revised model succeeded in fitting USAXS/SAXS and differently contrasted VSANS/SANS data of E. coli ATCC 25922 over four orders of magnitude in length scale. Specifically, this approach provides detailed insight into structural features of the cellular envelope, including the distance of the inner and outer membranes, as well as the scattering length densities of all bacterial compartments. The model was also successfully applied to E. coli K12, used for the authors' original modeling, as well as for two other E. coli strains. Significant differences were detected between the different strains in terms of bacterial size, intermembrane distance and its positional fluctuations. These findings corroborate the general applicability of the approach outlined here to quantitatively study the effect of bactericidal compounds on ultrastructural features of Gram-negative bacteria without the need to resort to any invasive staining or labeling agents.

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Increasing complexity in small-angle X-ray and neutron scattering experiments: from biological membrane mimics to live cells

Cited by 36 publications

References 110 publications

The effect of polydispersity, shape fluctuations and curvature on small unilamellar vesicle small-angle X-ray scattering curves

The effect of polydispersity, shape fluctuations and curvature on small unilamellar vesicle small-angle X-ray scattering curves

Changes in Membrane Protein Structural Biology

Evolution of the analytical scattering model of live Escherichia coli

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