Nordic Workshop on Scattering from Soft Matter

17 Jan 2018, 12:00 → 18 Jan 2018, 14:30 Europe/Stockholm

Auditorium (Medicon Village)

The 15th Nordic Workshop on Scattering from Soft Matter will be held in Lund, Sweden on January 17th to 18th, 2018.

There will be a full day program starting with lunch the first day and ending after lunch on the second day. The program includes invited and contributed talks, a poster session and a workshop dinner.

Registration is now closed

---

Important Note : Acceptance emails were sent out for oral and poster presentations before Christmas. A number of people have indicated that they did not recieve them. The abstracts accepted as oral presentations are those in the programme. If you requested an oral presentation and are not listed in the programme, then your abstract was accepted as a poster. If you requested a poster then your abstract was accepted.

---

Speakers and Programme

Confirmed keynote speakers include:

• Prof. Jan Skov Pedersen, Department of Chemistry and Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Denmark
• Dr. Hanna Isaksson, Biomedical Engineering, LU, Sweden
• Dr. Felix Roosen-Runge, Physical Chemistry, LU, Sweden
• Prof. Dr. Christian Gutt, Department of Physics, X-ray Physics, University of Siegen, Germany

The programme for the meeting can be found on the website here.

We look forward to seeing you in Lund in January 2018.

Support

The workshop is supported by:

LINXS - Lund Institute of Advanced Neutron and X-Ray Science (http://www.linxs.se)

Nordforsk Nordic Neutron Science Programme (https://www.nordforsk.org/en/programmes-and-projects/programmes/joint-nordic-programme-for-neutron-research) via the Nordic Network for Soft and Biological Matter.

Our corporate sponsors:

Anton Paar GmbH, Excillum AB, & Xenocs SA

  

And by MaxIV Laboratory, Lund University and the European Spallation Source

                                                                

Slides Book_of_Abstracts_NSSM2018.pdf

Wednesday, 17 January

12:00 → 13:00 Lunch

13:00 → 13:10 Opening of the conference and welcome

13:10 → 13:30 Overview of LINXS

Convener: Prof. Peter Schurtenberger (Lund University)

13:30 → 15:15 Session 1

13:30 Low radiation dose XPCS for dynamics in soft matter and biologial materials

X-ray radiation damage provides a serious bottle neck for investigating μs to s dynamics on nanometer length scales employing X-ray photon correlation spectroscopy. This limitation hinders the investigation of real time dynamics in most soft matter and biological materials which can tolerate only X-ray doses of kGy and below. Here, we show that this bottleneck can be overcome by low dose serial X-ray speckle visibility spectroscopy. The concept consists in spreading the dose needed for a correlation function over the entire sample volume by measuring at every spot on the sample the visibility of a speckle pattern as a function of exposure time. Mitigating the absorbed dose in this way is done at the expense of signal strength; the collected speckle patterns are sparse containing signal strength of 10-2 photons per pixel and possibly even less. We show that the speckle visibility correlation function can nevertheless be extracted by proper assignment of photon probabilities using a sufficiently large number of images. Employing X-ray doses of 640 Gy to 8.5 kGy and analyzing the sparse speckle patternswe follow as an example the slow nanoscale dynamics of an ionic liquid (IL) at the glass transition. Our method is especially relevant for the upcoming diffractiom limited storage rings providing a two orders of magnitude increase in coherent X-ray flux.

Speaker: Christian Gutt (University Siegen)

14:00 Structrural characterization of cellulose nanocrystals using SAXS

Cellulose is the main building block of trees and plants and the most abundant biopolymer in the world. It consists of highly ordered domains (nano crystals) and amorphous regions. These cellulose nanocrystals (CNC) have received significant interest due to their mechanical, optical, chemical, and rheological properties. CNC primarily obtained from naturally occurring cellulose fibers are biodegradable and renewable in nature and hence they serve as a sustainable and environmentally friendly material for most applications. These nanocrystals are basically hydrophilic in nature; however, they can be surface functionalized to meet various challenging requirements. The focus in this talk will be the structural characterization of CNC in different environments with small angle X-ray scattering (SAXS). It is employed to characterize the structure and shape of those crystallites directly after acid hydrolysis. Further, the rod-like crystallites are deposited and dried on different substrates and analysed using grazing incidence small angle X-ray scattering (GISAXS) and grazing incidence X-ray diffraction (GIXD).

Speaker: Dr Heike Ehmann (Anton Paar)

14:15 Short-time self-dynamics of immunoglobulin under bio- mimicking crowding conditions

Approximately 10-40% of the intra- and extracellular fluids of living organisms are occupied by macromolecules such as proteins. This macromolecular crowding condition was shown to influence reaction rates, and to lead to anomalous diffusion. The study of protein diffusion in such a crowded environment is, generally, not an easy task. Nevertheless, neutron backscattering (NBS) is well suited for measurements of the short-time self-diffusion of proteins in highly concentrated aqueous (D2O) solutions [1-4]. We present a NBS study on the pico- to nanosecond self-diffusion of the antibody proteins immunoglobulins (Ig) in aqueous environment. To systematically investigate the effect of macromolecular crowding on protein dynamics we vary the concentration of cellular lysate, mimicking a cellular environment. The dynamics of Ig in lysate is then compared with that of Ig in pure (heavy) water as a function of its own concentration (self-crowding) [2]. Despite the high polydispersity and the not easily predictable variance in lysate composition, the measured diffusion of Ig as a function of the overall volume fraction are in rather good agreement with those of Ig in the self-crowded environment at comparable volume fraction, suggesting a crucial role of hydrodynamic interactions and hence, in principle, the applicability of colloidal theories to model the protein short-time diffusion even in a cell-like environment. [1] Roosen-Runge F., Hennig M., Zhang F., Jacobs R.M.J., Sztucki M., Schober H., Seydel T., and Schreiber F. PNAS 108.29 (2011): 11815 [2] Grimaldo M., Roosen-Runge F., Zhang F., Seydel T., Schreiber F. JPCB 118.25 (2014): 7203. [3] Grimaldo M., Roosen-Runge F., Hennig M., Zanini F., Zhang F., Zamponi, M., Jalarvo N., Schreiber F., Seydel T. JPCL 6.13 (2015): 2577. [4] Braun M.K., Grimaldo M., Roosen-Runge F., Hoffmann I., Czakkel O., Sztucki M., Zhang F., Schreiber F., and Seydel T., JPCL 8.12 (2017): 2590.

Speaker: Dr Marco Grimaldo (ILL)

14:35 Interactions between Anionic Surfactants and Polymeric Micelles: stability and solubilisation kinetics

The kinetic processes involved in mixtures of surfactants and block copolymer micelles are not well understood. However, it is commonly known that surfactants exhibit rather fast equilibration kinetics, in the order of micro- to milliseconds, while polymers are much slower, in the order of minutes to months. In this contribution, we will present a study of the stability and solubilization kinetics of block copolymers micelles upon addition of sodium dodecyl sulphate (SDS) using small angle X-ray scattering (SAXS). We compare the ability of the surfactant to dissolve and form mixed micelles with two amphiphilic polymers; poly(ethylene propylene)-poly(ethylene oxide) (PEP-PEO) and end-capped PEO (C28-PEO). While the kinetics of C28PEO occurs on time scales on the order of minutes-hours on ambient temperatures, that of PEP1-PEO20 is known to be frozen on practical time scales. Addition of SDS to PEP1-PEO20 shows close to no change, even after extended period of time. However, upon addition of SDS to C28PEO5 we observe a fast dissolution and formation of mixed micelles, where the kinetics is seen to accelerate with the amount of added surfactant.

Speaker: Ms Synne Myhre (Department of Chemistry, University of Oslo)

14:55 End of Cooperativity: Chain Exchange Kinetics in Mixed Polymeric Micelles with Partially Crystalline Cores

Here we present a kinetic study on the chain exchange in mixed polymeric micelles containing partially crystalline cores. We are specifically interested in understanding how cooperative phenomena such as crystallization and melting affect the dynamics of self-assembled systems. As a model system we use n-alkyl-PEO ($C_{n}H_{2n+1}-O-(CH_{2}-CH_{2}-O)_{100}H$) with a molecular weight of roughly 5 kg/mol. In water these molecules form star-like micelles with a strongly segregated alkane core that partially crystallizes. This creates an additional energy barrier that needs to be overcome during chain expulsion.[1] We employ time-resolved small-angle neutron scattering in combination with the kinetic zero-average contrast technique to track the exchange kinetics.[2] We investigated mixtures of C28-PEO and C22-PEO and determined the respective melting enthalpies using differential scanning calorimetry (DSC) which was quantitatively compared to the kinetic data obtained from TR-SANS. We found that the core crystallization occurs cooperatively while the intermicellar chain exchange processes of C28-PEO and C22-PEO are virtually decoupled. Nevertheless the cooperative crystallization affects the separate exchange kinetics of both species. [1] Zinn, T., Willner, L., Pipich, V., Richter, D. and Lund, R. Effect of Core Crystallization and Conformational Entropy on the Molecular Exchange Kinetics of Polymeric Micelles. ACS Macro Letters 4, 651-655, doi:10.1021/acsmacrolett.5b00197 (2015). [2] Willner, L., Poppe, A., Allgaier, J., Monkenbusch, M. and Richter, D. Time-resolved SANS for the determination of unimer exchange kinetics in block copolymer micelles. Europhys. Lett. 55, 667-673, doi:DOI 10.1209/epl/i2001-00467-y (2001).

Speaker: Mr Nico König (Jülich Centre for Neutron Science JCNS and Institute for Complex Systems ICS, Forschungszentrum Jülich GmbH, 52425 Jülich, Germany AND Department of Chemistry, University of Oslo, P.O. Box 1033, Blindern, Oslo, Norway)

15:15 → 15:45 Coffee break

15:45 → 17:45 Session 2

15:45 Dynamics of Proteins in Solution Studied by Quasi-Elastic Neutron Scattering

Information on protein dynamics is of central importance for the understanding how biological function is effectuated by individual proteins as well as by well-adjusted interaction cascades within the crowded cytoplasm. Protein dynamics comprises a hierarchy of processes ranging from fluctuations of side chains and the backbone over interdomain motions to self-diffusion of the entire macromolecule and collective and cage diffusion characterizing the structural relaxation in crowded protein solutions. The broad distribution of time scales from pico- to microseconds, and the variety of dynamical processes including simple as well as confined, anomalous diffusion renders investigating protein dynamics a challenging research field. In this context, quasi-elastic neutron scattering (QENS) provides unique information on both the nature of the underlying dynamical process and the related geometry of dynamical confinement. Neutron backscattering (NBS) and neutron spin echo (NSE) spectroscopy have proven particularly relevant for proteins in solutions, as their instrumental time scales around nanoseconds allow to access simultaneously global and internal dynamics. After a brief overview on the key characteristics of QENS techniques, the scientific potential of QENS for protein dynamics will be examplified with two recent case studies. First, the changes of hierarchical protein dynamics upon thermal denaturation have been studied by both real-time monitoring and an additional detailed characterization of selected states [1,2]. Interestingly, while global dynamics are irreversibly arrested after denaturation, local internal dynamics change reversibly, suggesting that localized internal dynamics are mainly affected by basic physicochemical properties. Second, scenarios of dynamical arrest have been examined in solutions of α, β and γ crystallins as model systems for the eye lens with potential implications for the understanding of cataract and presbyopia. While α crystallin solutions behave similar to hard-sphere systems with a repulsive glass transition at high volume fraction, γB crystallin experiences a dramatic slowing down of cage diffusion already at comparably low volume fractions, suggesting an dynamical arrest driven by weak anisotropic attractions [3,4]. [1] M Grimaldo, F Roosen-Runge, et al. Phys. Chem. Chem. Phys. (2015) 17, 4645-4655 [2] M Hennig, F Roosen-Runge, et al. Soft Matter (2012) 8, 1628-1633 [3] S. Bucciarelli, J.S. Myung, et al. Sci. Adv. (2016) 2, e1601432 [4] S. Bucciarelli, L. Casal-Dujat, et al. J. Phys. Chem. Lett. (2015) 6, 4470–4474

Speaker: Felix Roosen-Runge (Division of Physical Chemistry, Lund University)

16:15 High throughput biological solution SAXS instrumentation for state-of-the-art structural biology research and validation.

Low volume samples, high throughput capabilities and easily reconfigurable instrument parameters for biological small angle x-ray scattering have previously been reserved for measurements at state-of-the-art synchrotron beamlines. With the introduction of the dedicated solution SAXS instrument, the BioXolver, Xenocs is not only moving the sample handling technology previously only seen at synchrotrons into the standard laboratory, but also providing it and the instrument itself with a level of automation that empower users at any skill level to obtain the best data for their particular sample without compromise. The instrument has been developed to be a truly easy to use and stable workhorse for samples in solution. By integrating computer vision technology, sample volumes down to 5 uL are possible and the in-line UV/VIS absorption measurements facilitate concentration estimation on the exposed sample. Automated sample loading, sample cell cleaning and drying is done with a high precision pipetting robot that also ensures gentle transport from the 2x96 well tray sample containers to the sample cell. No compromise is made on data quality as the detector is fully in vacuum, ensuring the lowest possible background. Furthermore, using a motorized detector stage and motorized scatterless slits, the sample to detector distance and flux can be automatically optimized to fit a large variety of protein complex sizes. Data reduction and analysis can also be automated and done using the open source software RAW that also includes integration and compatibility with the advanced software suite ATSAS from the EMBL. Built on the platform of an already successful instrument design, the BioXolver is an excellent choice for stability and data quality.

Speaker: Soren Skou (Xenocs)

16:30 Probing the structural dynamics of photoreceptor proteins by time-resolved X-ray solution scattering

The function of proteins is closely linked to their structure. Protein structures, however, are not static but rather dynamic and increased understanding of these structural dynamics is crucial for increased understanding of proteins. We have employed time-resolved X-ray solution scattering as a direct structural probe to study the signal transduction in different photoreceptor proteins. We have investigated members of the red-light sensing phytochromes, the blue light sensing cryptochromes and proteins with blue light sensing light-oxygen-voltage domains. Coupled with molecular dynamics simulations these investigations have revealed new information on how chemical changes at the light absorbing chromophore affects the global structure of the photoreceptor, in order to relay the signal further.

Speaker: Dr Oskar Berntsson (University of Gothenburg)

16:50 Probing the interaction between antimicrobial peptides and model biomembranes using small angle scattering techniques

Antibiotic resistance is one of the biggest threats to global health, according to the world health organization. Antimicrobial peptides (AMPs) is a group of molecules that are a natural part of the human immune system, shown to have effect against a broad spectrum of pathogens including both gram positive and gram negative bacteria.1 AMPs seem to be able to evade much of the bacterial resistance mechanisms, and are therefore promising candidates for future antibiotics. Instead of blocking specific biochemical pathways as most available antibiotic agents today, AMPs act physically on the cytoplasmic membrane itself. The precise microscopic mechanism for the perturbation of the membrane is not fully clear but several theories has been suggested including membrane deformation and pore forming which could lead to changes in the lipid dynamics, lateral and transversal composition and proton/ion transfer.2 Here we have used state of the art neutron and x-ray scattering techniques to investigate the microscopic mechanism of action of antimicrobial peptides with biomembranes as models for human and bacterial cell membranes. SAXS measurements on a model peptide, Indolicidin together with DMPC-DMPG-DMPE-PEG vesicles (with increasing amount of negatively charged DMPG) has shown that Indolicidin interacts with the model cell membrane causing a change in the contrast of the bilayer. Based on analysis of the results it seems that the peptide is situated at the interface between the lipid head group and the tail in the outer leaflet in the bilayer without significantly perturbing the structure of the bilayer. This is further supported by DSC where a shift and broadening of the melting lipid temperature upon addition of the peptide is observed. This indicates an associated disordering of the packing of the lipid tails in the bilayer of the vesicles. QCM-D experiments further confirms the disruption of the bilayer but also suggest removal of lipids upon flushing peptide solution over the bilayer surface. Preliminary SANS results do not indicate change in the lateral distribution of lipids which has been suggested as a possible mechanism on3). When combining the results from scattering methods, together with other complimentary techniques, we gain profound biophysical understanding of these systems. This knowledge can be used in the development of new antibiotics for the future based on antimicrobial peptides designed specifically for the task. 1. Fjell, C. D.; Hiss, J. A.; Hancock, R. E. W.; Schneider, G., Nat. Rev. Drug Discov. 2011, 11 (1), 37-51. 2. Nguyen, L. T.; Haney, E. F.; Vogel, H. J., Trends in biotechnology 2011, 29 (9), 464-472. 3. Epand, R. M.; Epand, R. F., Biochimica et Biophysica Acta (BBA)-Biomembranes 2009, 1788 (1), 289-294.

Speaker: Ms Josefine Eilsø Nielsen (University of Oslo)

17:10 Surface and colloid properties of oligomeric alkylglycosides

The increasing demand for environmentally friendly surfactants has resulted in a comprehensive research to identify surfactants that are biodegradable, non-toxic and produced from sustainable raw materials. In this respect, alkylglycosides with one or more sugar molecules, have shown not only promising surface properties but also favourable characteristics regarding sustainability, especially those with an oligomeric head group. However, the self-assembly and interfacial behaviour of oligomeric sugar surfactants are not very well understood. The aim with this study is therefore to get a deeper understanding of oligomeric alkylglycoside behaviour at interfaces and in solution in relation to their structure. The properties of different alkylglycosides, with focus on hexadecyl-β-D-maltopyranoside (C16G2) and a polydisperse mixture with tails of both 16 and 18 carbon atoms and head groups lengths between 1 and 20 glucose molecules (TZ30), have been characterized both in terms of interfacial (tensiometry, ellipsometry) and in bulk behaviour (DLS, SAXS, SANS, cryo-TEM). C16G2 has been found to form long wormlike micelles when dissolved in water above the cmc. The size of these micelles are relatively unaffected of temperature changes from 25 °C up to 90 °C and NaCl concentrations up to 1 M, while they are growing slightly with increasing concentration (0.5-10 mM). TZ30 are both forming small spherical micelles and other larger aggregates when dissolved in water at concentrations above the cmc. Ellipsometry measurements have shown that both C16G2 and TZ30 adsorbs at a hydrophobised silica surface to a higher extent than PEG-surfactants of comparable size.

Speaker: Mr Johan Larsson (Lund University)

17:30 Beamline Updates

17:45 → 19:00 Poster session

19:30 → 21:30 Conference dinner

Thursday, 18 January

08:30 → 10:00 Session 3

08:30 Scattering methods can unravel nano-structural changes in musculoskeletal tissues under loading

The musculoskeletal system enables locomotion of the human body, by force transfer through bone, cartilage, tendons and ligaments. Each tissue has a unique composition and hierarchical structure that result in an optimized mechanical function. Bone provides mechanical stability and support, while softer tissues such as tendons have a more damping function. With an aging population, the number of patients with musculoskeletal diseases, including fragile bones (osteoporosis), degenerated cartilage in the joints (osteoarthritis), and tendon pain (tendinopathy) is increasing. We use scattering methods as one tool to understand how the quality of musculoskeletal tissues are affected by age and disease. In combination with other high-resolution imaging methods, we unravel how composition, structure and orientation in these tissues are affected on the macro-, micro- and nano-scales. By studying the tissues in-situ, under concurrent mechanical loading, we can link the alteration in nanostructure to mechanical competence of the tissues, and thereby the anatomical function. As an example, we have combined experimental tensile testing inside SAXS and WAXS setups together with cameras on the surface and tomographic imaging, to study deformation simultaneous at multiple length scales in compact bone (Figure 1). We found that the orientation of the microstructure relative to the tensile loading influenced the strain magnitude on all length scales. Strains in the collagen fibers (measured by SAXS) were 2-3 times higher than the strains in the mineral crystals (measured by WAXS) for samples with microstructure oriented parallel to the loading. This will be extended to answer how the mineral crystal size and collagen fibre orientation and their response to load is altered in patients that are highly prone to bone fractures, e.g. osteoporosis. Alterations in tissue quality can be addressed by combining the knowledge from advanced structural, compositional and mechanical analyses. A better understanding of tissue quality help predicting the functional integrity of the tissue and supports the development of better diagnostics methods and treatment options. ![enter image description here][1] [1]: https://indico.esss.lu.se/event/874/picture/3.png Figure 1. Setup of experimental study to unravel the nanoscale behaviour in bone in different microstructural directions in situ during mechanical loading (Gustafsson et al., 2018).

Speaker: Hanna Isaksson (Lund University)

09:00 Structure Determination by SAXS​ in Protein Biophysics and Structural Biology

Jeppe LyngsøΔ, Eva Maria Steiner#, Jodie Guy#, Gleb Bourenkov¤, Ylva Lindqvist#, Thomas R. Schneider¤, Gunter Schneider#, Robert Schnell#, and Jan Skov PedersenΔ ΔDepartment of Chemistry and Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Gustav Wieds Vej 14, DK-8000 Aarhus, Denmark #Department of Medical Biochemistry and Biophysics, Karolinska Institutet, S-17 177 Stockholm, Sweden ¤Hamburg Unit c/o DESY, European Molecular Biology Laboratory (EMBL), Notkestrasse 85, 22603 Hamburg, Germany. The ATSAS package is probably the most widely used program package for analysis of SAXS data on biomolecules and biomolecular assemblies. However, in connection with recent work on several projects, we have experienced short-comings with the available programs. The most restrictive circumstance is that only executable programs are available, thus limiting the application to ‘standard’ projects that fall within the range of applications considered by the authors of the programs. For example, modelling protein-detergent complexes and inclusion of structure factors to describe inter-particle interactions are not possible within ATSAS. This together with a somewhat inconsistent description of hydration layers of composite structures, and the need for faster tools have motivated us to develop alternative programs. In this talk, we will present one of these projects as a case study, where the new rigid-body refinement methods have been developed and utilized with great success in parallel with the traditional approaches, so that comparisons are possible. A bacterial enzyme, involved in remodeling of periplasmic peptidoglycan structures, and its various recombinant constructs have been characterized by solution SAXS. Data from both SAXS instruments at Aarhus University have been utilized in the project. Probable solution structures were determined and verified by various approaches for the individual constructs based on known high-resolution structures of the domains and ab initio modelling. This contributed valuable information to the overall collaborative study on this interesting enzyme.

Speaker: Mr Jeppe Lyngsø (Aarhus University)

09:20 The Interaction of Perfluoroalkyl Substances with Mineral Surfaces and Biological Membranes

Perfluoroalkyl substances (PFASs) have hydrophobic and hydrophilic characteristics which makes them useful in many commercial and industrial applications such as textile, leather and paper impregnation, detergents, fire-fighting foam, etc. These compounds are receiving increasing global concern due to their persistence, bioaccumulation and possible adverse effects on the environment and living organisms [1]. In this project, we have investigated the interaction of PFASs with biological and mineral model system interfaces using neutron reflection. The PFAS were selected to allow comparing the effect of hydrophobic chain length and hydrophilic functional group on their interactions with the interface. The adsorption of PFASs was studied at two types of mineral surfaces (Al2O3, positively charged and SiO2, negatively charged). The electrostatic interaction was shown to be the driving force in the sorption process. The adsorbed PFAS could be removed by gentle rinsing with water. The adsorption process was shown to be influenced directly by the solubility limit of the PFAS which changes with the chain length [2]. Phospholipids are the building blocks of cell membranes and are commonly used as a model system to understand the fundamental behavior of biological membranes. DMPC (1,2-Dimyristoyl-sn-glycero-3-phosphocholine) bilayer was chosen as the model interface in this study. PFASs have shown to penetrate into the bilayer and displace lipids to accommodate themselves. Off-specular data from bilayers which have been immersed to PFASs indicate rough and patchy structures. Extensive rinsing with water can remove some PFAS, but a less dense bilayer is left behind. The interaction of these PFAS was shown to vary with both head group and chain length, and strongly correlate with the PFAS solubility limit [3]. 1. Hellsing et al. Chemosphere 159 (2016) 385e391. 2. Ahrens et al, Chemosphere 129 (2015) 33–38. 3. Nouhi et al, Journal of Colloid and Interface Science 155 (2018) 474-481.

Speaker: Ms Shirin Nouhi (Uppsala University)

09:40 Investigating the mesoscale of fibril hydrogels

Biomaterials widely presented in nature, like protein filaments in muscles, elastin in lungs, collagen in bones, cellulose fibers in fruits, and keratin in wool, self-assemble resulting in hierarchical structures that define their properties and uses. These hierarchical structures emerge from molecular and supra-molecular self-assembly systems organized on multiple length- and time-scales on the mesoscale. The study of such hierarchical structures at the mesoscale, where protein and carbohydrate self-assemblies like fibrils and tubes exist, is the particular interest of our research. Results from this study provide the knowledge to allow understanding the relationship between the architecture of fibril networks and their macroscopic properties, in order to connect the top (macroscale) to the bottom (microscale) and finally to understand how hierarchical systems, such as gels, are formed. This is achieved by measuring those properties using a combination of techniques; small-angle X-ray scattering for determining the molecular level self-assembly of the proteins and the resulting gel structure, cryo-scanning electron microscopy for investigating the 3D structural changes on the mesoscopic level, and bulk rheology for investigating the structural and kinetic changes on a macroscopic level.

Speaker: Dr Christina Efthymiou (Victoria University of Wellington, New Zealand and Uppsala University, Sweden)

10:00 → 10:30 Coffee break

10:40 → 12:15 Session 4

10:40 Refolding of SDS-unfolded proteins by non-ionic surfactants: Equilibrium and kinetics

Jan Skov Pedersen1,2, Jannik N. Pedersen1,2, Jeppe Lyngsø1,2, Jørn Døvling Kaspersen1,2, Anne Søndergaard1,2, Daniel Jhaf Madsen2, T. Zinn3, T. Narayanan3, and Daniel E. Otzen2 1Department of Chemistry, Aarhus University, Aarhus, Denmark 2Interdisciplinary Nanoscience center (iNANO), Aarhus University, Aarhus, Denmark 3European Synchrotron Radiation Facility, Grenoble, France email: jsp@chem.au.dk (J.S.P.) The strong and usually denaturing interaction between anionic surfactants (AS) and proteins/enzymes has both benefits and drawbacks: For example, it is in good use in electrophoretic mass determinations (SDS-PAGE) but limits enzyme efficiency in detergent formulations. Therefore, studies of the interactions between proteins and AS as well as non-ionic surfactants (NIS) are of both basic and applied relevance. The AS sodium dodecyl sulfate (SDS) denatures and unfolds globular proteins under most conditions. In contrast, it has been shown that the NIS octaethylene glycol monododecyl ether (C12E8) protects bovine serum albumin (BSA) from unfolding in SDS. We have shown recently that globular proteins unfolded by SDS can be refolded upon addition of C12E8. Four proteins, BSA, α-lactalbumin, (αLA), lysozyme (LYZ), and β-lactoglobulin (βLG), were studied by small-angle X-ray scattering (SAXS) and both near- and far-UV circular dichroism (CD). All proteins form complexes with SDS with a structural organization as protein-decorated micelles, in which the protein preserves secondary structure. For βLG, there is a quite spectacular transition from mainly -sheet structure in the native protein to mainly -helical structure in the complexes. All proteins were attempted refolded by the addition of C12E8. Except for apo αLA, which has a molten globular state, the proteins did not interact with C12E8 alone. The addition of C12E8 to the protein-SDS samples resulted, except for aLA, in refolding of the tested proteins and dissociation from surfactant micelles. It was concluded that C12E8 competes with globular proteins for association with SDS, making it possible to release and refold SDS-denatured proteins by adding sufficient amounts of C12E8. The last part of the talk will describe recent work using synchrotron radiation SAXS in combination with stopped-flow techniques on the kinetics of unfolding and refolding with emphasis on βLG. For this protein, the preliminary analysis shows a fast aggregation, when SDS is added, and a gradual conversion of the structure to highly symmetric protein-decorated micelle structures that is nearly complete in 10 s. The refolding is significantly slower with time constants of minutes, however, CD and Trp fluorescence in our home lab reveal that there is also a much faster process that is not captured in the SAXS measurements, which is a secondary structure conversion. In the work, both simple analysis in terms of measured basis functions (SAXS data) as well as modelling on absolute scale have been used and will be briefly described. Kaspersen JD, Søndergaard A, Madsen DJ, Otzen DE, Pedersen JS. Refolding of SDS-Unfolded Proteins by Nonionic Surfactants. Biophys J. 2017 112(8):1609-1620.

Speaker: Jan Skov Pedersen (1Department of Chemistry, Aarhus University, Aarhus, Denmark)

11:00 Liquid-Metal-Jet X-ray Source for In-situ SAXS studies in the Home Laboratory

High-end x-ray scattering techniques such as SAXS, BIO-SAXS, non-ambient SAXS and GISAXS rely heavily on the x-ray source brightness for resolution and exposure time. Traditional solid or rotating anode x-ray tubes are typically limited in brightness by when the e-beam power density melts the anode. The liquid-metal-jet technology has overcome this limitation by using an anode that is already in the molten state. We have previously demonstrated prototype performance of a metal-jet anode x-ray source concept with unprecedented brightness in the range of one order of magnitude above current state-of-the art sources. Over the last years, the liquid-metal-jet technology has developed from prototypes into fully operational and stable X-ray tubes running in many labs over the world. Small angle scattering has been identified as a key application for this x-ray tube technology, since this application benefits greatly from high-brightness and small spot-sizes, to achieve a high flux x-ray beam with low divergence. Multiple users and system manufacturers have since installed the metal-jet anode x-ray source into their SAXS set-ups with successful results. With the high brightness from the liquid-metal-jet x-ray source, in-situ SAXS studies can be performed – even in the home laboratory. The influence of the size of the x-ray source and its distance to the x-ray optics on the divergence will be discussed, and how to minimize the divergence and maximize the flux in SAXS experiments targeted to specific applications. This presentation will review the current status of the metal-jet technology specifically in terms of stability, lifetime, flux and brightness. It will also discuss details of the liquid-metal-jet technology with a focus on the fundamental limitations of the technology. It will furthermore refer to some recent SAXS and GISAXS data from users of metal-jet x-ray tubes.

Speaker: Shichao Hu (Excillum)

11:15 Time-resolved SAXS reveals ionic liquid interaction with model membranes

Ionic liquids (ILs) are solvents that have many desirable properties. For example, they are easier to handle than volatile organic solvents and they dissolve cellulose. However, many are harmful. The toxicity of ILs is in part due to their effects on cell membranes. In this study, we have observed effects of (tetradecyl)tributylphosphonium acetate ([P$_{14444}$][OAc]), trioctylmethylphosphonium acetate ([P$_{8881}$][OAc]), tributylmethylphosphonium acetate ([P$_{4441}$][OAc]) and 1-ethyl-3-methylimidazolium acetate ([emim][OAc]) on model membranes composed of L-α-phosphatidylcholine multilamellar vesicles (MLV) by time-resolved small-angle X-ray scattering (SAXS). The SAXS experiments were conducted on beamline ID02 at ESRF, Grenoble. The penetration of ILs into MLVs was studied at millisecond resolution using a stopped-flow mixing device. The results show that all studied ILs penetrated the lipid bilayers. [emim][OAc] and [P$_{4441}$][OAc] caused a thinning of the lamellar distance but did not induce disorder in the lamellae. The ILs with larger cations destroy the lamellar order. The results give insight into the ways in which ILs can penetrate the cell membrane and cause changes to it.

Speaker: Dr Inkeri Kontro (University of Helsinki)

11:35 Characterization of liposomal formulations to treat Fabry Disease

Fabry disease is a lysosomal storage disorder, where the lack of α-Galactosidase A (GLA) causes accumulation of glycosphingolipids leading to damage of the kidneys, heart and nervous system. With current treatments, free GLA is injected intravenously in patients causing instability of GLA, high immunogenicity, and low bioavailability. To overcome this, liposomes has been developed that can encapsulate GLA and improve the performance. Further development of the liposomes is necessary and currently being done to archive perfect control of the assembly process. The liposomes consists of several different components that can alter the size, polydispersity and structure of the liposomes, and characterization of different formulations is therefore important in the development of new liposomal formulations. We have used an In-house small-angle X-ray scattering (SAXS) setup to get information on the average liposomal structure in presence of different lipids. We use a para crystalline model (Guinier, A. (1963). X-ray Diffraction, San Francisco: Freeman) with a finite number of layers and disorder between layers, while a set of Gaussians is used to describe the cross-section profile of the lipid bilayer. Scattering from GLA contributes to the total signal and is in the model added to the scattering from the bilayers, so that an estimate of the amount of GLA can be obtained. Using the model for fitting the data, we are able to follow incorporation of GLA in the liposomes and changes in bilayer thickness as well as changes in the amount of multi-lamellar structures and bilayer ordering. The incorporation of GLA in the formulations correlates well with the results from the SAXS data, and it is also observed that the distance between layers is increased when larger molecules are incorporated in the liposomes. The number of bilayers determined with SAXS correlates well with that seen with cryo-TEM. Altogether, the SAXS data can give us a lot of information on the structure, composition and polydispersity of the liposomes. We also used static and dynamic light scattering to obtain information on the size and polydispersity of the samples and can use this in characterizing the homogeneity of the samples. This information is important for further development of new liposomal drug formulations and will be an important tool in the design of liposomes that has a low polydispersity, are stable and can help in improving the treatment of Fabry Disease. This project has received funding from the European Union’s Horizon 2020 research and innovation programmer under the grant agreement No 720942.

Speaker: Mr Jannik Pedersen (Department of Chemistry and Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Gustav Wieds Vej 14, 8000 Aarhus C, Denmark)

11:55 Inverse- and real-space scattering of aqueous diblock copolymer micelles

Poly(glycerol monomethacrylate)–poly(benzyl methacrylate) (PGMA–PBzMA) diblock copolymer micelles were synthesized via polymerization-induced self-assembly (PISA) using reversible additional–fragmentation chain-transfer (RAFT) aqueous emulsion polymerization in D$_2$O. PISA reactions produce polymer nanoparticles in situ during the reaction and can be performed at high particle concentrations. PGMA–PBzMA synthesized by PISA is known to form only spherical micelles, making it an ideal model system for exploring new characterization methods. The structure of the polymer micelles was obtained using small-angle X-ray scattering (SAXS) and, a more recently developed form of neutron scattering, spin-echo small-angle neutron scattering (SESANS). Fitting the scattering data from these techniques showed that the inverse-space (SAXS) and real-space (SESANS) scattering gave structural parameters that compare very favorably. As far as we are aware, this is the first report of polymer micelles being studied by SESANS. Using both an inverse-space and a real-space scattering technique, with optimal sensitivity at different lengthscales, has made it possible to gain interesting information about both the structure of and interactions between polymer micelles as a dilute dispersion (SAXS) and directly in the synthesis medium (SESANS).

Speaker: Dr Smith Gregory (Københavns Universitet)

12:15 → 12:30 Sum up and end of conference

12:30 → 13:30 Lunch