Zinc mediates control of nitrogen fixation via transcription factor filamentation.

Plants adapt to fluctuating environmental conditions by adjusting their metabolism and gene expression to maintain fitness1. In legumes, nitrogen homeostasis is maintained by balancing nitrogen acquired from soil resources with nitrogen fixation by symbiotic bacteria in root nodules2-8. Here we show that zinc, an essential plant micronutrient, acts as an intracellular second messenger that connects environmental changes to transcription factor control of metabolic activity in root nodules. We identify a transcriptional regulator, FIXATION UNDER NITRATE (FUN), which acts as a sensor, with zinc controlling the transition between an inactive filamentous megastructure and an active transcriptional regulator. Lower zinc concentrations in the nodule, which we show occur in response to higher levels of soil nitrate, dissociates the filament and activates FUN. FUN then directly targets multiple pathways to initiate breakdown of the nodule. The zinc-dependent filamentation mechanism thus establishes a concentration readout to adapt nodule function to the environmental nitrogen conditions. In a wider perspective, these results have implications for understanding the roles of metal ions in integration of environmental signals with plant development and optimizing delivery of fixed nitrogen in legume crops.

A Targetable N-Terminal Motif Orchestrates α-Synuclein Oligomer-to-Fibril Conversion.

Oligomeric species populated during α-synuclein aggregation are considered key drivers of neurodegeneration in Parkinson's disease. However, the development of oligomer-targeting therapeutics is constrained by our limited knowledge of their structure and the molecular determinants driving their conversion to fibrils. Phenol-soluble modulin α3 (PSMα3) is a nanomolar peptide binder of α-synuclein oligomers that inhibits aggregation by blocking oligomer-to-fibril conversion. Here, we investigate the binding of PSMα3 to α-synuclein oligomers to discover the mechanistic basis of this protective activity. We find that PSMα3 selectively targets an α-synuclein N-terminal motif (residues 36-61) that populates a distinct conformation in the mono- and oligomeric states. This α-synuclein region plays a pivotal role in oligomer-to-fibril conversion as its absence renders the central NAC domain insufficient to prompt this structural transition. The hereditary mutation G51D, associated with early onset Parkinson's disease, causes a conformational fluctuation in this region, leading to delayed oligomer-to-fibril conversion and an accumulation of oligomers that are resistant to remodeling by molecular chaperones. Overall, our findings unveil a new targetable region in α-synuclein oligomers, advance our comprehension of oligomer-to-amyloid fibril conversion, and reveal a new facet of α-synuclein pathogenic mutations.

Poly(Sitosterol)-Based Hydrophobic Blocks in Amphiphilic Block Copolymers for the Assembly of Hybrid Vesicles.

Amphiphilic block copolymer and lipids can be assembled into hybrid vesicles (HVs), which are an alternative to liposomes and polymersomes. Block copolymers that have either poly(sitostryl methacrylate) or statistical copolymers of sitosteryl methacrylate and butyl methacrylate as the hydrophobic part and a poly(carboxyethyl acrylate) hydrophilic segment are synthesized and characterized. These block copolymers assemble into small HVs with soybean L-α-phosphatidylcholine (soyPC), confirmed by electron microscopy and small-angle X-ray scattering. The membrane's hybrid nature is illustrated by fluorescence resonance energy transfer between labeled building blocks. The membrane packing, derived from spectra when using Laurdan as an environmentally sensitive fluorescent probe, is comparable between small HVs and the corresponding liposomes with molecular sitosterol, although the former show indications of transmembrane asymmetry. Giant HVs with homogenous distribution of the block copolymers and soyPC in their membranes are assembled using the electroformation method. The lateral diffusion of both building blocks is slowed down in giant HVs with higher block copolymer content, but their permeability toward (6)-carboxy-X-rhodamine is higher compared to giant vesicles made of soyPC and molecular sitosterol. This fundamental effort contributes to the rapidly expanding understanding of the integration of natural membrane constituents with designed synthetic compounds to form hybrid membranes.

Living with epilepsy in adolescence and young adulthood transitioning from pediatric to adult hospital services: A systematic review and meta-synthesis of qualitative studies.

BACKGROUND: Transition is characterized by developing greater self-identity and growing independence, but adolescents dealing with chronic illnesses encounter health-related and situational changes during transition. Despite the many suggestions made in recent years, the shift from pediatric to adult care continues to pose difficulties for adolescents and young adults with epilepsy (AWE). The holistic perspective of AWE's experiences and needs during transition is not as well understood. AIM: To synthesize the qualitative evidence related to AWE's experiences and needs transitioning from pediatric to adult hospital care. METHODS: This systematic review adhered to the rigorous Joanna Briggs methodology for qualitative evidence synthesis. A comprehensive search was conducted across multiple databases, including PubMed, CINAHL, Scopus, Embase, PsycINFO, and ProQuest Dissertations & Theses Global, from their inception to April 2024. The findings were critically appraised and aggregated using meta-synthesis. RESULTS: The search yielded a total of 3,985 studies, and twenty-one were included in the review. Two of the included studies were undertaken in a program where a transition clinic was established. The meta-synthesis reveals that the transition experience of AWE is more than a change from one clinic to another and is interwoven into a pattern of developmental, health-illness, situational, and organizational transition issues. Five synthesized findings were developed: 1) Feeling different from others and striving to address the impact of epilepsy in everyday life; 2) the transition from pediatric to adult care - a problematic intersection point; 3) the family's role - support or parental overprotectiveness 4) seeking knowledge and being familiar with epilepsy supported by healthcare professionals and technologies, and 5) development of independence and responsibility through involvement and support from healthcare professionals and parents. CONCLUSION: During the transition from pediatric to adult hospital care, AWEs encounter a loss of familiarity, increased responsibility, and feelings of not belonging. Therefore, it is essential to create an environment where they can thrive beyond the limitations of their illness. Understanding, acceptance, and inclusivity should characterize this environment to support AWEs in facilitating the development of responsibility, independence, and confidence as they navigate transitions.

Periostin C-Terminal Is Intrinsically Disordered and Interacts with 143 Proteins in an In Vitro Epidermal Model of Atopic Dermatitis.

Human periostin is a 78-91 kDa matricellular protein implicated in extracellular matrix remodeling, tumor development, metastasis, and inflammatory diseases like atopic dermatitis, psoriasis, and asthma. The protein consists of six domains, including an N-terminal Cys-rich CROPT domain, four fasciclin-1 domains, and a C-terminal domain. The exons encoding the C-terminal domain may be alternatively spliced by shuffling four exons, generating ten variants of unknown function. Here, we investigate the structure and interactome of the full-length variant of the C-terminal domain with no exons spliced out. The structural analysis showed that the C-terminal domain lacked a tertiary structure and was intrinsically disordered. In addition, we show that the motif responsible for heparin-binding is in the conserved very C-terminal part of periostin. Pull-down confirmed three known interaction partners and identified an additional 140 proteins, among which nine previously have been implicated in atopic dermatitis. Based on our findings, we suggest that the C-terminal domain of periostin facilitates interactions between connective tissue components in concert with the four fasciclin domains.

Induction, inhibition, and incorporation: Different roles for anionic and zwitterionic lysolipids in the fibrillation of the functional amyloid FapC.

Amyloid proteins are widespread in nature both as pathological species involved in several diseases and as functional entities that can provide protection and storage for the organism. Lipids have been found in amyloid deposits from various amyloid diseases and have been shown to strongly affect the formation and structure of both pathological and functional amyloid proteins. Here, we investigate how fibrillation of the functional amyloid FapC from Pseudomonas is affected by two lysolipids, the zwitterionic lipid 1-myristoyl-2-hydroxy-sn-glycero-3-phosphocholine and the anionic lipid 1-myristoyl-2-hydroxy-sn-glycero-3-phospho-(1'-rac-glycerol) (LPG). Small-angle X-ray scattering, circular dichroism, dynamic light scattering, and thioflavin T fluorescence measurements were performed simultaneously on the same sample to ensure reproducibility and allow a multimethod integrated analysis. We found that LPG strongly induces fibrillation around its critical micelle concentration (cmc) by promoting formation of large structures, which mature via accumulation of intermediate fibril structures with a large cross section. At concentrations above its cmc, LPG strongly inhibits fibrillation by locking FapC in a core-shell complex. In contrast, lipid 1-myristoyl-2-hydroxy-sn-glycero-3-phosphocholine induces fibrillation at concentrations above its cmc, not via strong interactions with FapC but by being incorporated during fibrillation and likely stabilizing the fibrillation nucleus to reduce the lag phase. Finally, we show that LPG is not incorporated into the fibril during assembly but rather can coat the final fibril. We conclude that lipids affect both the mechanism and outcome of fibrillation of functional amyloid, highlighting a role for lipid concentration and composition in the onset and mechanism of fibrillation in vivo.

Position statement on the definition, incidence, diagnosis and outcome of acute on chronic pancreatitis.

BACKGROUND: Acute on chronic pancreatitis (ACP) is a relatively common condition, but there are significant gaps in our knowledge on the definition, incidence, diagnosis, treatment and prognosis. METHODS: A systematic review that followed PICO (Population; Intervention; Comparator; Outcome) recommendation for quantitative questions and PICo (Population, Phenomenon of Interest, Context) for qualitative research was done to answer 10 of the most relevant questions about ACP. Quality of evidence was judged by the GRADE criteria (Grades of Recommendation, Assessment, Development and Evaluation). The manuscript was sent for review to 12 international experts from various disciplines and continents using a Delphi process. RESULTS: The quality of evidence, for most statements, was low to very low, which means that the recommendations in general are only conditional. Despite that, it was possible to reach strong levels of agreement by the expert panel for all 10 questions. A new consensus definition of ACP was reached. Although common, the real incidence of ACP is not known, with alcohol as a major risk factor. Although pain dominates, other non-specific symptoms and signs can be present. Serum levels of pancreatic enzymes may be less than 3 times the upper limit of normal and cross-sectional imaging is considered more accurate for the diagnosis in many cases. It appears that it is less severe and with a lower mortality risk than acute pancreatitis. CONCLUSIONS: Although the evidence base is poor, this position statement provides a foundation from which to advance management of ACP.

Human myelin proteolipid protein structure and lipid bilayer stacking.

The 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.

Structural Investigations of Human A2M Identify a Hollow Native Conformation That Underlies Its Distinctive Protease-Trapping Mechanism.

Human α2-macroglobulin (A2M) is the most characterized protease inhibitor in the alpha-macroglobulin (αM) superfamily, but the structure of its native conformation has not been determined. Here, we combined negative stain electron microscopy (EM), small-angle X-ray scattering (SAXS), and cross-linking-mass spectrometry (XL-MS) to investigate native A2M and its collapsed conformations that are obtained through aminolysis of its thiol ester by methylamine or cleavage of its bait region by trypsin. The combined interpretation of these data resulted in a model of the native A2M tetramer and its conformational changes. Native A2M consists of two crescent-shaped disulfide-bridged subunit dimers, which face toward each other and surround a central hollow space. In native A2M, interactions across the disulfide-bridged dimers are minimal, with a single major interface between the linker (LNK) regions of oppositely positioned subunits. Bait region cleavage induces both intrasubunit domain repositioning and an altered configuration of the disulfide-bridged dimer. These changes collapse the tetramer into a more compact conformation, which encloses an interior protease-trapping cavity. A recombinant A2M with a modified bait region was used to map the bait region's position in native A2M by XL-MS. A second recombinant A2M introduced an intersubunit disulfide into the LNK region, demonstrating the predicted interactions between these regions in native A2M. Altogether, our native A2M model provides a structural foundation for understanding A2M's protease-trapping mechanism, its conformation-dependent receptor interactions, and the dissociation of native A2M into dimers due to inflammatory oxidative stress.

Mutation-induced dimerization of transforming growth factor-ß-induced protein may drive protein aggregation in granular corneal dystrophy.

Protein aggregation in the outermost layers of the cornea, which can lead to cloudy vision and in severe cases blindness, is linked to mutations in the extracellular matrix protein transforming growth factor-ß-induced protein (TGFBIp). Among the most frequent pathogenic mutations are R124H and R555W, both associated with granular corneal dystrophy (GCD) characterized by the early-onset formation of amorphous aggregates. The molecular mechanisms of protein aggregation in GCD are largely unknown. In this study, we determined the crystal structures of R124H, R555W, and the lattice corneal dystrophy-associated A546T. Although there were no changes in the monomeric TGFBIp structure of any mutant that would explain their propensity to aggregate, R124H and R555W demonstrated a new dimer interface in the crystal packing, which is not present in wildtype TGFBIp or A546T. This interface, as seen in both the R124H and R555W structures, involves residue 124 of the first TGFBIp molecule and 555 in the second. The interface is not permitted by the Arg124 and Arg555 residues of wildtype TGFBIp and may play a central role in the aggregation exhibited by R124H and R555W in vivo. Using cross-linking mass spectrometry and in-line size exclusion chromatography-small-angle X-ray scattering, we characterized a dimer formed by wildtype and mutant TGFBIps in solution. Dimerization in solution also involves interactions between the N- and C-terminal domains of two TGFBIp molecules but was not identical to the crystal packing dimerization. TGFBIp-targeted interventions that disrupt the R124H/R555W crystal packing dimer interface might offer new therapeutic opportunities to treat patients with GCD.

Structural Basis for Dityrosine-Mediated Inhibition of α-Synuclein Fibrillization.

α-Synuclein (α-Syn) is an intrinsically disordered protein which self-assembles into highly organized ß-sheet structures that accumulate in plaques in brains of Parkinson's disease patients. Oxidative stress influences α-Syn structure and self-assembly; however, the basis for this remains unclear. Here we characterize the chemical and physical effects of mild oxidation on monomeric α-Syn and its aggregation. Using a combination of biophysical methods, small-angle X-ray scattering, and native ion mobility mass spectrometry, we find that oxidation leads to formation of intramolecular dityrosine cross-linkages and a compaction of the α-Syn monomer by a factor of √2. Oxidation-induced compaction is shown to inhibit ordered self-assembly and amyloid formation by steric hindrance, suggesting an important role of mild oxidation in preventing amyloid formation.

A model on an absolute scale for the small-angle X-ray scattering from bovine casein micelles.

Casein micelles extracted from milk are 100-400 nm-sized particles, made up of proteins and calcium phosphates, with the latter as colloidal calcium phosphate particles (CCPs) in a size range of 2-4 nm embedded in a protein network. The hierarchical structures give rise to a variation of scattering intensity over many orders of magnitude, which can be measured by small-angle X-ray scattering and static light scattering. Expressions for the scattering intensity of a general simple model for composite particles with polydispersities of overall size and subparticles are derived, and some approximations are checked by generating scattering data for systems generated by Monte Carlo simulations. Based on the simpler models, a new model has been developed for casein micelles, where the scattering is expressed on an absolute scale and where the concentrations of, respectively, protein and CCPs are used as constraints, providing a consistent model. The CCPs are modelled as oblate ellipsoids and the protein as star structures. Correlations between the substructures of CCPs and protein structures are taken into account in terms of partial structure factors. The overall structure as well as some heterogeneities at intermediate length scale are modelled as polydisperse spheres. The model fits the data very well on all length scales and demonstrates that both the scattering from CCPs and protein is important. Thus, the model provides a detailed description of the casein structure, which is consistent with the information available in the literature.

Confusion with the definition and diagnostic criteria for acute on chronic pancreatitis: review and recommendations.

OBJECTIVES: Chronic pancreatitis (CP) is a fibroinflammatory disease complicated by episodes of acute inflammation (acute on chronic pancreatitis (ACP)). This entity is common, variably defined and can reflect different pathological mechanisms that requires different interventions. The aim of this study is to conduct a systematic review of how ACP is described, defined and diagnosed in the literature. METHODS: A systematic search was conducted from January 1993 to June 2020. All articles that used a term to describe ACP in adults were reviewed and definitions and diagnostic criteria were sought. RESULTS: After reviewing 2271 abstracts, 848 articles included a term to describe ACP. The most common descriptions were 'acute on/in CP' (374), 'acute exacerbation of CP' (345) and 'flare(-up) of CP' (43). Among the 848 articles, 14 included a pragmatic definition of ACP, and only 2 papers stated diagnostic criteria. These covered both acute inflammation and acute exacerbation of chronic abdominal pain. CONCLUSION: There is no universally accepted term, definition or diagnostic criteria for ACP. A consensus definition is needed to improve quality and comparability of future articles as well as clinical management.

Universal effective interactions of globular proteins close to liquid-liquid phase separation: Corresponding-states behavior reflected in the structure factor.

Intermolecular interactions in protein solutions, in general, contain many contributions. If short-range attractions dominate, the state diagram exhibits liquid-liquid phase separation (LLPS) that is metastable with respect to crystallization. In this case, the extended law of corresponding states (ELCS) suggests that thermodynamic properties are insensitive to details of the underlying interaction potential. Using lysozyme solutions, we investigate the applicability of the ELCS to the static structure factor and how far effective colloidal interaction models can help to rationalize the phase behavior and interactions of protein solutions in the vicinity of the LLPS binodal. The (effective) structure factor has been determined by small-angle x-ray scattering. It can be described by Baxter's adhesive hard-sphere model, which implies a single fit parameter from which the normalized second virial coefficient b2 is inferred and found to quantitatively agree with previous results from static light scattering. The b2 values are independent of protein concentration but systematically vary with temperature and solution composition, i.e., salt and additive content. If plotted as a function of temperature normalized by the critical temperature, the values of b2 follow a universal behavior. These findings validate the applicability of the ELCS to globular protein solutions and indicate that the ELCS can also be reflected in the structure factor.

Improving brain B0 shimming using an easy and accessible multi-coil shim array at ultra-high field.

OBJECT: Improve shimming capabilities of ultra-high field systems, with addition of an accessible low-complexity B0 shim array for head MRI at 7 T. MATERIALS AND METHODS: An eight channel B0 shim coil array was designed as a tradeoff between shimming improvement and construction complexity, to provide an easy to use shim array that can be employed with the standard 7 T head coil. The array was interfaced using an open-source eight-channel shim amplifier rack. Improvements in field homogeneity for whole-brain and slice-based shimming were compared to standard second-order shimming, and to more complex higher order dynamic shimming and shim arrays with 32 and 48 channels. RESULTS: The eight-channel shim array provided 12% improvement in whole brain static shimming and provided 33% improvement when using slice-based shimming. With this, the eight-channel array performed similar to third-order dynamic shimming (without the need for higher order eddy current compensation). More complex shim arrays with 32 and 48 channels performed better, but require a dedicated RF coil. DISCUSSION: The designed eight-channel shim array provides a low-complexity and low-cost approach for improving B0 field shimming on an ultra-high field system. In both static and dynamic shimming, it provides improved B0 homogeneity over standard shimming.

A multimethod approach for analyzing FapC fibrillation and determining mass per length.

Amyloid proteins are found in a wide range of organisms owing to the high stability of the ß-sheet core of the amyloid fibrils. There are both pathological amyloids involved in various diseases and functional amyloids that play a beneficial role for the organism. The aggregation process is complex and often involves many different species. Full understanding of this process requires parallel acquisition of data by complementary techniques monitoring the time course of aggregation. This is not an easy task, given the often-stochastic nature of aggregation, which can lead to significant variations in lag time. Here, we investigate the aggregation process of the functional amyloid FapC by simultaneous use of four different techniques, namely dynamic light scattering, small-angle x-ray scattering (SAXS), circular dichroism, and Thioflavin T fluorescence. All these approaches are applied to the same FapC sample just after desalting. Our data allow us to construct a master time-course graph showing the same time-course of aggregation by all techniques. This allows us to integrate insights from approaches that report on different structural and length scales. During the lag phase, loosely aggregated oligomers with random-coil structure are formed, which subsequently transform to fibrils without accumulation of additional significant species. Subsequently, the loosely associated protofilaments/subfilaments, which form side by side, mature to more compact fibrils. Furthermore, we determine the mass per length of the mature fibrils, obtaining very similar results by SAXS (33 kDa/nm) and tilted-beam transmission electron microscopy (31 kDa/nm). Transmission electron microscopy showed that the fibrils consist of primarily two protofilaments and similar dimensions of the cross section of the fibrils as revealed by SAXS modeling when the number of protofilaments per fibril was taken into account. Mass per length information underscores the general usefulness of SAXS in fibrillation analysis and provides an important constraint for further modeling the fibril structures.

Cys-labeling kinetics of membrane protein GlpG: a role for specific SDS binding and micelle changes?

Empirically, α-helical membrane protein folding stability in surfactant micelles can be tuned by varying the mole fraction MFSDS of anionic (sodium dodecyl sulfate (SDS)) relative to nonionic (e.g., dodecyl maltoside (DDM)) surfactant, but we lack a satisfying physical explanation of this phenomenon. Cysteine labeling (CL) has thus far only been used to study the topology of membrane proteins, not their stability or folding behavior. Here, we use CL to investigate membrane protein folding in mixed DDM-SDS micelles. Labeling kinetics of the intramembrane protease GlpG are consistent with simple two-state unfolding-and-exchange rates for seven single-Cys GlpG variants over most of the explored MFSDS range, along with exchange from the native state at low MFSDS (which inconveniently precludes measurement of unfolding kinetics under native conditions). However, for two mutants, labeling rates decline with MFSDS at 0-0.2 MFSDS (i.e., native conditions). Thus, an increase in MFSDS seems to be a protective factor for these two positions, but not for the five others. We propose different scenarios to explain this and find the most plausible ones to involve preferential binding of SDS monomers to the site of CL (based on computational simulations) along with changes in size and shape of the mixed micelle with changing MFSDS (based on SAXS studies). These nonlinear impacts on protein stability highlights a multifaceted role for SDS in membrane protein denaturation, involving both direct interactions of monomeric SDS and changes in micelle size and shape along with the general effects on protein stability of changes in micelle composition.

Investigation of the enhanced ability of bile salt surfactants to solubilize phospholipid bilayers and form mixed micelles.

The self-assembly in mixtures of the anionic bile salt surfactant sodium deoxycholate (NaDC) and the zwitterionic phospholipid 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) in physiological saline solution has been investigated using light scattering, small-angle X-ray scattering and cryo-transmission electron microscopy. Rather small tri-axial ellipsoidal NaDC-DMPC mixed micelles form at a high content of bile salt in the mixture, which increase in size as an increasing amount of DMPC is incorporated into the micelles. Eventually, the micelles begin to grow substantially in length to form long wormlike micelles. At higher mole fractions of DMPC, the samples become turbid and cryo-TEM measurements reveal the existence of large perforated vesicles (stomatosomes), coexisting with geometrically open disks. To our knowledge, stomatosomes have not been observed before for any bile salt-phospholipid system. Mixed micelles are found to be the sole aggregate structure in a very wide regime of bile salt-phospholipid compositions, i.e. up to about 77 mol% phospholipid in the micelles. This is much higher than the corresponding value of 25 mol% observed for the conventional surfactant hexadecyltrimethylammonium bromide (CTAB) mixed with DMPC in the same solvent. The enhanced ability of bile salt surfactants to solubilize phospholipid bilayers and form mixed micelles is rationalized using bending elasticity theory. From our theoretical analysis, we are able to conclude that amphiphilic molecules rank in the following order of increasing spontaneous curvature: phospholipids < conventional surfactants < bile salts. The bending rigidity of the different amphiphilic molecules increases according to the following sequence: bile salts < conventional surfactants < phospholipids.

Interactions in protein solutions close to liquid-liquid phase separation: ethanol reduces attractions via changes of the dielectric solution properties.

Ethanol is a common protein crystallization agent, precipitant, and denaturant, but also alters the dielectric properties of solutions. While ethanol-induced unfolding is largely ascribed to its hydrophobic parts, its effect on protein phase separation and inter-protein interactions remains poorly understood. Here, the effects of ethanol and NaCl on the phase behavior and interactions of protein solutions are studied in terms of the metastable liquid-liquid phase separation (LLPS) and the second virial coefficient B2 using lysozyme solutions. Determination of the phase diagrams shows that the cloud-point temperatures are reduced and raised by the addition of ethanol and salt, respectively. The observed trends can be explained using the extended law of corresponding states as changes of B2. The results for B2 agree quantitatively with those of static light scattering and small-angle X-ray scattering experiments. Furthermore, B2 values calculated based on inter-protein interactions described by the Derjaguin-Landau-Verwey-Overbeek (DLVO) potential and considering the dielectric solution properties and electrostatic screening due to the ethanol and salt content quantitatively agree with the experimentally observed B2 values.

Toward reliable low-density lipoprotein ultrastructure prediction in clinical conditions: A small-angle X-ray scattering study on individuals with normal and high triglyceride serum levels.

Atherosclerosis is the main killer in the west and therefore a major health challenge today. Total serum cholesterol and lipoprotein concentrations, used as clinical markers, fail to predict the majority of cases, especially between the risk scale extremes, due to the high complexity in lipoprotein structure and composition. In particular, low-density lipoprotein (LDL) plays a key role in atherosclerosis development, with LDL size being a parameter considered for determining the risk for cardiovascular diseases. Determining LDL size and structural parameters is challenging to address experimentally under physiological-like conditions. This article describes the biochemistry and ultrastructure of normolipidemic and hypertriglyceridemic LDL fractions and subfractions using small-angle X-ray scattering. Our results conclude that LDL particles of hypertriglyceridemic compared to healthy individuals 1) have lower LDL core melting temperature, 2) have lower cholesteryl ester ordering in their core, 3) are smaller, rounder and more spherical below melting temperature, and 4) their protein-containing shell is thinner above melting temperature.