Browsing by Author "Darwish, TA"
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- ItemAdvancements in the provision of deuterated lipids for neutron applications from the National Deuteration Facility(Australian Institute of Nuclear Science and Engineering (AINSE), 2020-11-11) Yepuri, NR; Moir, M; Krause-Heuer, AM; Klenner, MA; Darwish, TAMolecular deuteration significantly increases the options for structure-function investigations using neutron scattering and diffraction techniques. Chemical deuteration activities, where catalysed 1H/2H exchange is followed by custom chemical synthesis, have led to diverse neutron scattering and reflectometry studies previously hampered by the lack of appropriate scattering contrast in multi-component samples. Deuteration of phospholipids is a common practice to elucidate membrane structure, dynamics and function, by providing selective visualisation in neutron scattering. Although analogous deuterium? (2H) and hydrogen? containing (1H) molecules have similar physicochemical properties, these isotopes of hydrogen result in vastly different for neutron scattering signals. Over the past few years the National Deuteration Facility (ANSTO) has increased its synthetic capability to produce complex deuterated molecules including lipids and phospholipids. Such synthetically challenging molecules are perdeuterated phytantriol1, tail deuterated POPC, and perdeuterated POPC.2 Phytantriol is an interfacially-active lipid that is chemically robust, non-digestible and forms particles with internal bicontinuous cubic phase structures (cubosomes) when dispersed with non-ionic surfactants at physiological temperatures. The tail-deuterated POPC, perdeuterated POPC and tail-deuterated GMO isotopologues would also provide suitable contrast for many neutron experiments and so these have been also our synthetic targets. Recently neutron reflection was employed to investigate the impact of phospholipid saturation (POPC-d64) and presence of cholesterol in cell model membranes on LDL and HDL lipid exchange and removal processes.3 Neutron reflection data that distinguish the effect of phospholipid acyl chain saturation and the presence of cholesterol on the ability of lipoproteins to exchange lipids to/from model membrane will be presented.
- ItemANSTO’s National Deuteration Facility: recent advancements and an overview on molecular deuteration capabilities for neutron applications(Australian Institute of Nuclear Science and Engineering (AINSE), 2020-11-11) Wilde, KL; Cagnes, MP; Duff, AP; Klenner, MA; Krause-Heuer, AM; Moir, M; Rekas, A; Russell, RA; Yepuri, NR; Darwish, TAThe National Deuteration Facility (NDF) at the Australian Nuclear Science and Technology Organisation (ANSTO) provides deuteration through both biological and chemical techniques for a diversity of molecules and applications and is the only facility of its type in the Southern Hemisphere with the specialised expertise and infrastructure for both biological and chemical molecular deuteration. Molecular deuteration of organic compounds and biomolecules significantly increases the options in complex structure function investigations using neutron scattering and reflectometry, nuclear magnetic resonance (NMR), mass spectrometry (MS) and other techniques. Deuteration (substitution of the naturally occurring hydrogen stable isotope deuterium (2H or D) for 1H (or H)) can provide contrast and improved resolution to assist investigations into the relationship between molecular structure and function of molecules of both biological and synthetic origin. By developing a suite of capabilities in both in vivo deuteration of biomolecules and chemical deuteration of small organic molecules, the NDF provides access to a broad range of deuterated molecules for research and industry. Variably deuterated proteins can be produced via recombinant expression in Escherichia coli and other microbial systems utilised to produce deuterated cellulose and cholesterol. By tailoring deuteration approaches with the ongoing development of chemical deuteration protocols for a broader range of molecular classes than available commercially, the NDF has increased the range of systems that can be investigated using deuterated molecules. Lipids, phospholipids (including head or tail or head/tail deuterated mono-unsaturated lipids such as POPC and DOPC), heterocyclics, aromatics, surfactants, ionic liquids, saturated and unsaturated fatty acids, sugars and match-out detergents have been deuterated. Common neutron applications include partially deuterated proteins for SANS experiments investigating multiprotein systems, neutron crystallography of perdeuterated proteins, neutron reflectometry of lipid bilayers systems and SANS using saturated lipid vesicles, or detergents amongst others. An overview and update on the NDF will be provided which will include details on the NDF User Program (e.g. information on the available modes of access), recent advancements in custom deuterated molecules available and brief highlights of deuterated molecule utilisation for neutron experiments at ANSTO’s Australian Centre for Neutron Scattering (ACNS). © 2020 The Authors.
- ItemApolipoprotein E binding drives structural and compositional rearrangement of mRNA-containing lipid nanoparticles(American Chemical Society (ACS), 2021-03-23) Sebastiani, F; Yanez Arteta, M; Lerche, M; Porcar, L; Lang, C; Bragg, RA; Elmore, CS; Krishnamurthy, VR; Russell, RA; Darwish, TA; Pichler, H; Waldie, S; Moulin, M; Haertlein, M; Forsyth, VT; Lindfors, L; Cárdenas, MEmerging therapeutic treatments based on the production of proteins by delivering mRNA have become increasingly important in recent times. While lipid nanoparticles (LNPs) are approved vehicles for small interfering RNA delivery, there are still challenges to use this formulation for mRNA delivery. LNPs are typically a mixture of a cationic lipid, distearoylphosphatidylcholine (DSPC), cholesterol, and a PEG-lipid. The structural characterization of mRNA-containing LNPs (mRNA-LNPs) is crucial for a full understanding of the way in which they function, but this information alone is not enough to predict their fate upon entering the bloodstream. The biodistribution and cellular uptake of LNPs are affected by their surface composition as well as by the extracellular proteins present at the site of LNP administration, e.g., apolipoproteinE (ApoE). ApoE, being responsible for fat transport in the body, plays a key role in the LNP's plasma circulation time. In this work, we use small-angle neutron scattering, together with selective lipid, cholesterol, and solvent deuteration, to elucidate the structure of the LNP and the distribution of the lipid components in the absence and the presence of ApoE. While DSPC and cholesterol are found to be enriched at the surface of the LNPs in buffer, binding of ApoE induces a redistribution of the lipids at the shell and the core, which also impacts the LNP internal structure, causing release of mRNA. The rearrangement of LNP components upon ApoE incubation is discussed in terms of potential relevance to LNP endosomal escape. © 2021 American Chemical Society. Open Access. This publication is licensed under CC-BY 4.0.
- ItemThe Australian National Deuteration Facility for structure function applications using neutrons(International Conference on Neutron Scattering, 2017-07-12) Darwish, TA; Yepuri, NR; Heuer, AK; Duff, AP; Wilde, KL; Holden, PJThere have been limited global initiatives in the field of molecular deuteration where the majority of these programs focus on biological deuteration of proteins and lipids, while more complex deuterated small molecules haven’t been widely available to the neutron community. This has limited the experiments that can be performed, and formed a bottle-neck for advancing the applications of neutron scattering. In this paper we will discuss the recent advancements and the impact of deuteration on the research outcomes achieved by using deuterated molecules produced by the National Deuteration Facility of the Australian Nuclear Science and Technology Organisation. Recent high-impact case studies in the fields of molecular electronics, structural biology, and biotechnology will be presented which reveal the exciting and diverse characterisation studies which are now available for the neutron community.
- ItemBehaviour of single transmembrane peptides during in meso crystallization from the contrast-matched lipidic cubic phase of monoolein(Australian Institute of Nuclear Science and Engineering (AINSE), 2018-11-19) van't Hag, L; de Campo, L; Tran, N; Sokolova, AV; Trenker, R; Call, M; Garvey, CJ; Leung., A; Darwish, TA; Krause-Heuer, AM; Knott, RB; Meikle, T; Gras, S; Drummond, CJ; Mezzenga, R; Conn, CIn meso membrane protein crystallization within a lipidic mesophase has revolutionized the structural biology of integral membrane proteins (IMPs). High-resolution structures of these proteins are crucial to understanding fundamental cellular processes at a molecular level, and can lead to new and improved treatments for a wide range of diseases via rational drug design. However, overall success rates of the promising in meso crystallization technique remain low because of a fundamental lack of understanding about factors that promote crystal growth. In particular, to date, two decades from invention of the method, the protein-eye-view of the in meso crystallization mechanism had not been solved. We have investigated this for the first time using small-angle neutron scattering (SANS). Contrast-matching between the scattering of the lipid membrane formed by MO and the aqueous solution was used to isolate and track the scattering of single-transmembrane peptides during the growth of protein crystals in meso. No peptide enrichment was observed at the flat points of the diamond cubic QIID phase of MO in contrast to suggestions in several modeling studies. During in meso crystallization of the DAP12 peptide a decrease in form factor and a transient fluid lamellar Lα phase could be observed providing direct evidence for the proposed crystallization mechanism. Synthesis of fully deuterated MO was required for this purpose and scattering of this new material in various solvents and under a range of conditions will be described, specifically regarding the effect of the relative scattering length densities (SLD) of the headgroup, acyl chain and solvent, which can advance the use of neutron scattering with other self-assembly materials. © The Authors.
- ItemBiopolymer deuteration for neutron scattering and other isotope-sensitive techniques(Elsevier, 2015) Russell, RA; Garvey, CJ; Darwish, TA; Foster, LJR; Holden, PJThe use of microbial biosynthesis to produced deuterated recombinant proteins is a well-established practice in investigations of the relationship between molecular structure and function using neutron scattering and nuclear magnetic resonance spectroscopy. However, there have been few reports of using microbial synthetic capacity to produce labeled native biopolymers. Here, we describe methods for the production of deuterated polyhydroxyalkanoate biopolyesters in bacteria, the polysaccharide chitosan in the yeast Pichia pastoris, and cellulose in the bacterium Gluconacetobacter xylinus. The resulting molecules offer not only multiple options in creating structural contrast in polymer blends and composites in structural studies but also insight into the biosynthetic pathways themselves. © 2015 Elsevier Inc.
- ItemCellulose dissolution in ionic liquid: ion binding revealed by neutron scattering(ACS Publications, 2018-09-20) Raghuwanshi, VS; Cohen, Y; Garnier, GFG; Garvey, CJ; Russell, RA; Darwish, TA; Garnier, GDissolution of cellulose in 1-ethyl-3-methylimidazolium acetate (EMIMAc) ionic liquid (IL) was investigated by small-angle neutron scattering (SANS) with contrast variation. Cellulose and EMIMAc of different deuteration levels provide sufficient contrast in revealing the cellulose dissolution processes. Two experiments were performed: hydrogenated microcrystalline cellulose (MCC) was dissolved in deuterated IL (IL-D14), and deuterated bacterial cellulose (DBC) was dissolved in hydrogenated IL (IL-H14). Contrary to the expectation of high contrast between MCC and IL-D14, a dramatic reduction of the measured intensity (scattering cross section) was observed, about 1/3 of the value predicted based on the scattering length density (SLD) difference. This is attributed to the tight binding of acetate ions to the cellulose chains, which reduces the SLD difference. Measurements using small-angle X-ray scattering (SAXS) corroborate this effect by indicating increased contrast due to ion adsorption resulting in enhanced SLD difference. The experiments performed with DBC dissolution in IL-H14 suggest the presence of fractal aggregates of the dissolved cellulose, indicating lower solubility compared to the MCC. Contrast variation SANS measurements highlight tight ion binding of at least one acetate ion per anhydroglucose unit (AGU). EMIMAc is a successful cellulose solvent, as in addition to disrupting intermolecular hydrogen bonding, it imparts effective charge to the cellulose chains hindering their agglomeration in solution. © 2018 American Chemical Society
- ItemCharacterizing the photoinduced switching process of a nitrospiropyran self-assembled monolayer using in situ sum frequency generation spectroscopy(American Chemical Society, 2012-10-02) Darwish, TA; Tong, YJ; James, M; Hanley, TL; Peng, QL; Ye, SSum frequency generation (SFG) vibrational spectroscopy is employed to investigate the reversible, photoinduced spiro -> merocyanine isomerization of a self-assembled monolayer, the result of attachment of nitrospiropyran to a gold surface using a dithiolane anchoring group. The attachment of these molecular "alligator clips" to spiropyran molecules provide an easily accessible method to self-assemble a robust monolayer of spiropyran on a gold surface, which allows photoswitching of the spiropyran units. Probing the symmetric and antisymmetric stretching modes of the nitro group allows the determination of the structural orientation of the charged moiety with respect to the surface normal as well as the isomerization rates under photoinduced switching conditions. The photoisomerization of the spiropyran SAM on the gold surface is much faster than the rates of switching spiropyrans in a solid crystalline form, and the rate of thermal relaxation of the opened to closed form in this study is found to be on the same time scale as the relaxation of spiropyran when present in solutions with polar solvents. © 2012, American Chemical Society.
- ItemChemical deuteration and neutrons for structure function applications(Australian Institute of Nuclear Science and Engineering, 2016-11-29) Darwish, TA; Yepuri, NR; Heuer, AK; Cagnes, MP; Holden, PJIn small angle neutron scattering and neutron reflectometry studies, the use of mixtures of deuterated and hydrogenated solvents to manipulate scattering length density and achieve contrast variation is widespread. This approach, while useful is less effective for multicomponent organic systems containing molecules of similar scattering length densities. In such systems molecular deuteration is necessary to achieve contrast between the different components and it significantly increases the options in structure function investigations. There have been limited global initiatives in the field of molecular deuteration where the majority of these programs focus on biological deuteration of proteins and lipids, while more complex deuterated small molecules haven’t been widely available to the neutron community. This has limited the experiments that can be performed, and formed a bottle neck for advancing the applications of neutron scattering. In this paper we will discuss the recent advancements and the impact of deuteration on the research outcomes achieved by using deuterated molecules produced by the chemical deuteration laboratories at ANSTO’s National Deuteration Facility. Recent high-impact case studies will be presented which reveal the exciting and diverse characterisation studies which are now available for the neutron community. The chemical deuteration of surfactants, sugars, heterocyclic and aromatic compounds has made possible a wide range of investigations systems in the fields of molecular electronics, structural biology, and biotechnology.
- ItemChemical deuteration of ionic liquids and their application in neutron reflectivity(International Conference on Neutron Scattering, 2017-07-12) Akutsu, K; Darwish, TA; Cagnes, MP; Tamura, K; Kanaya, TLow melting imidazolium salts are one of the most popular and widely used ionic liquids (ILs). Due to their low vapor pressures and thermal stability, ILs have been widely studied and employed in many applications such as synthesis and catalysis solvents, extraction solvents, electrolyte, and biopolymer solvents. However, in many case, structural analysis of IL samples is difficult because the X-ray and neutron scattering contrast between the ILs and the organic solute is not enough to allow different elements to be clearly discerned. Partial or complete deuteration of organic compounds for contrast variation in the scattering length densities of materials is one of the most effective techniques in the application of neutron scattering analysis. Therefore, this technique is ideally suited for the structural analysis of IL samples using neutron scattering techniques. In this study, we synthesized several kinds of deuterated 1-alkyl-3-methylimidazolium ionic liquids (alkyl = ethyl, butyl, and octyl) for studying the electric double layer structure in ILs using neutron reflectivity technique. For this purpose, we have developed a simple and effective method for the deuteration of imidazolium salts ILs on gram scale, starting from their protonated versions. In this presentation, we will show the summary of deuteration experiments and the results of neutron reflectivity analysis of the electric double layer structure in ILs.
- ItemChemical disorder in a frustrated J1/J2 quantum spin chain material(Australian Institute of Physics, 2018-01-30) Rule, KC; Mole, RA; Zanardo, J; Krause-Heuer, AM; Darwish, TA; Lerch, MLFRecently a new one-dimensional (1D) quantum spin chain system has been synthesised: catena-dichloro(2-Cl-3Mpy)copper(II), [where 2-Cl-3Mpy=2-chloro-3-methylpyridine]. We shall refer to this compound as cd-Cu. Preliminary calculations and bulk magnetic property measurements indicate that this system does not undergo magnetic ordering down to 1.8K and is a prime candidate for investigating frustration in a J1/J2 system (where the nearest neighbour interactions, J1, are ferromagnetic and the next nearest neighbour interactions, J2, are antiferromagnetic) [1]. Calculations predicted 3 possible magnetic interaction strengths for J1 below 6meV depending on the orientation of the ligand [2]. For one of the predicted J1values, the existence of a quantum critical point is implied. A deuterated sample of cd-Cu was produced at the National Deuteration Facility and the excitations measured using the PELICAN TOF spectrometer. Scattering was weak from this sample, but indicated the most likely scenario involves an average of the 3 possible magnetic excitations in this material, rather than the random array of exchange interactions as predicted by Herringer et al., [2]. This may indicate the possibility of tuning the chemical structure to favour a system which may exhibit a quantum critical point.
- ItemCO2 triggering and controlling orthogonally multiresponsive photochromic systems(American Chemical Society, 2010-08-11) Darwish, TA; Evans, RA; James, M; Malic, N; Triani, G; Hanley, TLWe report a new generic method of reversibly controlling the photochromism of spiropyrans. It was found that the photochromic effect of spiropyrans can be reversibly switched on and off by addition and removal of carbon dioxide (CO2) to spiropyran in alcohol solutions containing an amidine (i.e., DBU) that acts as a CO2 sensitizer. Spiropyrans are not photochromic in the presence of DBU but photochromic when CO2 is subsequently added to the solution. The CO2 is readily removed by inert gas bubbling, thus allowing facile activation and deactivation of the photochromic effect. Carbon dioxide, without the presence of the sensitizing amidine, had no effect on photochromism of the spiropyrans. Other photochromic dyes classes such as spirooxazines and chromenes are not affected by this CO2/DBU stimulus. As a result, orthogonal activation of mixtures of spirooxazines and spiropyrans was achieved to provide four color states (clear, yellow, green, and blue) by varying the combinations of the stimuli of UV, visible light, CO2, and CO2 depleted. This finding now permits the many applications using spiropyrans to be CO2 responsive. © 2010, American Chemical Society
- ItemComprehensive study of carbon dioxide adsorption in the metal–organic frameworks M2(dobdc) (M = Mg, Mn, Fe, Co, Ni, Cu, Zn)(Royal Society of Chemistry, 2014-08-28) Queen, WL; Hudson, MR; Bloch, ED; Mason, JA; Gonzalez, MI; Lee, JS; Gygi, D; Howe, JD; Lee, K; Darwish, TA; James, M; Peterson, VK; Teat, SJ; Smit, B; Neaton, JB; Long, JR; Brown, CMAnalysis of the CO2 adsorption properties of a well-known series of metal–organic frameworks M2(dobdc) (dobdc4− = 2,5-dioxido-1,4-benzenedicarboxylate; M = Mg, Mn, Fe, Co, Ni, Cu, and Zn) is carried out in tandem with in situ structural studies to identify the host–guest interactions that lead to significant differences in isosteric heats of CO2 adsorption. Neutron and X-ray powder diffraction and single crystal X-ray diffraction experiments are used to unveil the site-specific binding properties of CO2 within many of these materials while systematically varying both the amount of CO2 and the temperature. Unlike previous studies, we show that CO2 adsorbed at the metal cations exhibits intramolecular angles with minimal deviations from 180°, a finding that indicates a strongly electrostatic and physisorptive interaction with the framework surface and sheds more light on the ongoing discussion regarding whether CO2 adsorbs in a linear or nonlinear geometry. This has important implications for proposals that have been made to utilize these materials for the activation and chemical conversion of CO2. For the weaker CO2 adsorbents, significant elongation of the metal–O(CO2) distances are observed and diffraction experiments additionally reveal that secondary CO2 adsorption sites, while likely stabilized by the population of the primary adsorption sites, significantly contribute to adsorption behavior at ambient temperature. Density functional theory calculations including van der Waals dispersion quantitatively corroborate and rationalize observations regarding intramolecular CO2 angles and trends in relative geometric properties and heats of adsorption in the M2(dobdc)–CO2 adducts. © 2014, The Royal Society of Chemistry.
- ItemDeuterated phospholipids to study the structure, function and dynamics of membrane proteins using neutron scattering(Australian Nuclear Science and Technology Organisation, 2021-11-26) Yepuri, NR; Moir, M; Krause-Heuer, AM; Darwish, TAContrast matching and contrast variation in neutron scattering provide unparalleled power for understanding the structure, function, and dynamics of a selected component in a multicomponent system. A sophisticated contrast study often requires the availability of deuterated molecules in which deuterium atoms are introduced in a predictable and controlled fashion to replace protons. This can be achieved by direct deuteration of precursors followed by custom chemical synthesis, for which expertise and capabilities have been developed at facility (NDF), ANSTO. It this paper we will discuss recent high impact research output using deuterated phospholipids produced by NDF/ANSTO. We will describe the synthesis and applications of selectively or perdeuterated unsaturated phospholipids to contrast match out the whole lipid bilayer or nano disks within a multicomponent system. Further, we also describe their role in investigations related to membrane lipoproteins (ApoE) exchange in relation to lipid unsaturation,[1] effect of membrane composition,[2] and conformational analysis Mg+2 channel by neutron scattering techniques.[2, 3]
- ItemDeuterated polymers for probing phase separation using infrared microspectroscopy(ACS Publications, 2013-12-23) Russell, RA; Darwish, TA; Puskar, L; Martin, DE; Holden, PJ; Foster, LJRInfrared (IR) microspectroscopy has the capacity to determine the extent of phase separation in polymer blends. However, a major limitation in the use of this technique has been its reliance on overlapping peaks in the IR spectra to differentiate between polymers of similar chemical compositions in blends. The objective of this study was to evaluate the suitability of deuteration of one mixture component to separate infrared (IR) absorption bands and provide image contrast in phase separated materials. Deuteration of poly(3-hydroxyoctanoate) (PHO) was achieved via microbial biosynthesis using deuterated substrates, and the characteristic C–D stretching vibrations provided distinct signals completely separated from the C–H signals of protonated poly(3-hydroxybutyrate) (PHB). Phase separation was observed in 50:50 (% w/w) blends as domains up to 100 μm through the film cross sections, consistent with earlier reports of phase separation observed by scanning electron microscopy (SEM) of freeze-fractured protonated polymer blends. The presence of deuterated phases throughout the film suggests there is some miscibility at smaller length scales, which increased with increasing PHB content. These investigations indicate that biodeuteration combined with IR microspectroscopy represents a useful tool for mapping the phase behavior of polymer blends.
- ItemDeuterated squalene and sterols from modified Saccharomyces cerevisiae(Royal Society of Chemistry (RSC), 2023-07-31) Recsei, C; Russell, RA; Cagnes, MP; Darwish, TAUniformly deuterated sterols and biosynthetically related materials are important for neutron, NMR, tracing and bioanalysis studies as well as critical tools for the creation of improved lipid nanoparticle formulations. The production of sufficient quantities of materials relies not only on the engineering of microorganisms to selectively accumulate desired materials but also methods for the isolation, purification and characterisation of these materials to ensure their usefulness. Uniformly deuterated squalene, the universal precursor to sterols in biological systems, has been produced and characterised. Cholesterol has been produced with controlled levels of uniform deuteration, increased biosynthetic yield and a methodology developed for the extraction and purification of this material without HPLC. Two sterols, not previously produced in deuterated forms, have been prepared with uniform deuteration: 22,23-dihydrobrassicasterol and 24-methylenecholesterol. This report triples the number of sterols that have been produced with uniform deuteration, purified and characterised and provides a silylation/silver ion chromatography protocol for the separation of sterols which differ by the degree of unsaturation. The techniques for the 13C NMR analysis of deuterated sterols, site-specific deuteration levels and an analysis of key biosynthetic steps based on these data are reported. © 2023 The Royal Society of Chemistry, Open Access, published under a CC-BY-NC licence.
- ItemDeuteration for biological SANS: case studies, success and challenges in chemistry and biology(Elsevier, 2022-11) Duff, AP; Cagnes, MP; Darwish, TA; Krause-Heuer, AM; Moir, M; Recsei, C; Rekas, A; Russell, RA; Wilde, KL; Yepuri, NRSmall angle neutron scattering is a powerful complementary technique in structural biology. It generally requires, or benefits from, deuteration to achieve its unique potentials. Molecular deuteration has become a mature expertise, with deuteration facilities located worldwide to support access to the technique for a wide breadth of structural biology and life sciences. The sorts of problems well answered by small angle scattering and deuteration involve large (> 10 Å) scale flexible movements, and this approach is best used where high-resolution methods (crystallography, NMR, cryo-EM) leave questions unanswered. This chapter introduces deuteration, reviewing biological deuteration of proteins, lipids and sterols, and then steps through the ever-expanding range of deuterated molecules being produced by chemical synthesis and enabling sophisticated experiments using physiologically relevant lipids. Case studies of recent successful use of deuteration may provide illustrative examples for strategies for future experiments. We discuss issues of nomenclature for synthesised molecules of novel labeling and make recommendations for their naming. We reflect on our experiences, with cost associated with achieving an arbitrary deuteration level, and on the benefits of experimental co-design by user scientist, deuteration scientist, and neutron scattering scientist working together. Although methods for biological and chemical deuteration are published in the public domain, we recommend that the best method to deuterate is to engage with a deuteration facility. © 2022 Elsevier
- ItemFluorine-18 radiolabelling and in vitro / in vivo metabolism of [18F]D4-PBR111(John Wiley & Sons, Inc, 2019-05-26) Wyatt, NA; Safavi-Naeini, M; Wotherspoon, ATL; Arthur, A; Nguyen, AP; Parmar, A; Hamze, H; Day, CM; Zahra, D; Matesic, L; Davis, E; Rahardjo, GL; Yepuri, NR; Shepherd, R; Murphy, RB; Pham, TQ; Nguyen, VH; Callaghan, PD; Holden, PJ; Grégoire, MC; Darwish, TA; Fraser, BHObjectives The purinergic receptor P2X ligand-gated ion channel type 7 (P2X7R) is an adenosine triphosphate (ATP)-gated ion-channel, and P2X7R is a key player in inflammation. P2X7R is an emerging therapeutic target in central nervous system (CNS) diseases including Alzheimer's disease (AD) and Parkinson's disease (PD), because P2X7R also plays a pivotal role in neuroinflammation. P2X7R represents a potential molecular imaging target for neuroinflammation via biomedical imaging technique positron emission tomography (PET), and several radioligands targeting P2X7R have been developed and evaluated in animals. In our previous work, we have developed and characterized [11C]GSK1482160 as a P2X7R radioligand for neuroinflammation,2 clinical evaluation of [11C]GSK1482160 in healthy controls and patients is currently underway, and the estimation of radiation dosimetry for [11C]GSK1482160 in normal human subjects has been reported.3 Since the half-life (t1/2) of radionuclide carbon-11 is only 20.4 min, it is attractive for us to develop derivatives of [11C]GSK1482160, which can be labeled with the radionuclide fluorine-18 (t1/2, 109.7 min), and a fluorine-18 ligand would be ideal for widespread use.4 To this end, a series of [18F]fluoroalkyl including [18F]fluoromethyl (FM), [18F]fluoroethyl (FE), and [18F]fluoropropyl (FP) derivatives of GSK1482160 have been prepared and examined as new potential P2X7R radioligands. © 2019 The Authors
- ItemHigh-density lipoprotein function is modulated by the SARS-CoV-2 spike protein in a lipid-type dependent manner(Elsevier B. V., 2023-09) Correa, YB; Del Giuduce, R; Waldie, S; Thépaut, M; Gerelli, Y; Moulin, M; Delauney, C; Fieschi, F; Haertlein, M; Le Brun, AP; Forsyth, VT; Moir, M; Russell, RA; Darwish, TA; Brinck, J; Wodaje, T; Jansen, M; Martín, C; Roosen-Runge, F; Cárdenas, M; Micciulla, S; Pichler, HThere is a close relationship between the SARS-CoV-2 virus and lipoproteins, in particular high-density lipoprotein (HDL). The severity of the coronavirus disease 2019 (COVID-19) is inversely correlated with HDL plasma levels. It is known that the SARS-CoV-2 spike (S) protein binds the HDL particle, probably depleting it of lipids and altering HDL function. Based on neutron reflectometry (NR) and the ability of HDL to efflux cholesterol from macrophages, we confirm these observations and further identify the preference of the S protein for specific lipids and the consequent effects on HDL function on lipid exchange ability. Moreover, the effect of the S protein on HDL function differs depending on the individuals lipid serum profile. Contrasting trends were observed for individuals presenting low triglycerides/high cholesterol serum levels (LTHC) compared to high triglycerides/high cholesterol (HTHC) or low triglycerides/low cholesterol serum levels (LTLC). Collectively, these results suggest that the S protein interacts with the HDL particle and, depending on the lipid profile of the infected individual, it impairs its function during COVID-19 infection, causing an imbalance in lipid metabolism. © Crown Copyright 2023. Published by Elsevier Inc. Open Access - CC BY licence 4.0.
- ItemHuman myelin proteolipid protein structure and lipid bilayer stacking(Springer Nature, 2022-07-12) Ruskamo, S; Raasakka, A; Pedersen, JS; Martel, A; Škubník, K; Darwish, TA; Porcar, L; Kursula, PThe myelin sheath is an essential, multilayered membrane structure that insulates axons, enabling the rapid transmission of nerve impulses. The tetraspan myelin proteolipid protein (PLP) is the most abundant protein of compact myelin in the central nervous system (CNS). The integral membrane protein PLP adheres myelin membranes together and enhances the compaction of myelin, having a fundamental role in myelin stability and axonal support. PLP is linked to severe CNS neuropathies, including inherited Pelizaeus-Merzbacher disease and spastic paraplegia type 2, as well as multiple sclerosis. Nevertheless, the structure, lipid interaction properties, and membrane organization mechanisms of PLP have remained unidentified. We expressed, purified, and structurally characterized human PLP and its shorter isoform DM20. Synchrotron radiation circular dichroism spectroscopy and small-angle X-ray and neutron scattering revealed a dimeric, α-helical conformation for both PLP and DM20 in detergent complexes, and pinpoint structural variations between the isoforms and their influence on protein function. In phosphatidylcholine membranes, reconstituted PLP and DM20 spontaneously induced formation of multilamellar myelin-like membrane assemblies. Cholesterol and sphingomyelin enhanced the membrane organization but were not crucial for membrane stacking. Electron cryomicroscopy, atomic force microscopy, and X-ray diffraction experiments for membrane-embedded PLP/DM20 illustrated effective membrane stacking and ordered organization of membrane assemblies with a repeat distance in line with CNS myelin. Our results shed light on the 3D structure of myelin PLP and DM20, their structure–function differences, as well as fundamental protein–lipid interplay in CNS compact myelin. © 2022 The Authors - Open Access under a Creative Commons Attribution 4.0.
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