Single Condensate Research Articles

Spatiotemporal formation of a single liquid-like condensate and amyloid fibrils of α-synuclein by optical trapping at solution surface

Liquid-like protein condensates have recently attracted much attention due to their critical roles in biological phenomena. They typically show high fluidity and reversibility for exhibiting biological functions, while occasionally serving as sites for the formation of amyloid fibrils. To comprehend the properties of protein condensates that underlie biological function and pathogenesis, it is crucial to study them at the single-condensate level; however, this is currently challenging due to a lack of applicable methods. Here, we demonstrate that optical trapping is capable of inducing the formation of a single liquid-like condensate of α-synuclein in a spatiotemporally controlled manner. The irradiation of tightly focused near-infrared laser at an air/solution interface formed a condensate under conditions coexisting with polyethylene glycol. The fluorescent dye-labeled imaging showed that the optically induced condensate has a gradient of protein concentration from the center to the edge, suggesting that it is fabricated through optical pumping-up of the α-synuclein clusters and the expansion along the interface. Furthermore, Raman spectroscopy and thioflavin T fluorescence analysis revealed that continuous laser irradiation induces structural transition of protein molecules inside the condensate to β-sheet rich structure, ultimately leading to the condensate deformation and furthermore, the formation of amyloid fibrils. These observations indicate that optical trapping is a powerful technique for examining the microscopic mechanisms of condensate appearance and growth, and furthermore, subsequent aging leading to amyloid fibril formation.

Multi-condensate state as a functional strategy to optimize the cell signaling output

The existence of multiple biomolecular condensates inside living cells is a peculiar phenomenon not compatible with the predictions of equilibrium statistical mechanics. In this work, we address the problem of multiple condensates state (MCS) from a functional perspective. We combine Langevin dynamics, reaction-diffusion simulation, and dynamical systems theory to demonstrate that MCS can indeed be a function optimization strategy. Using Arp2/3 mediated actin nucleation pathway as an example, we show that actin polymerization is maximum at an optimal number of condensates. For a fixed amount of Arp2/3, MCS produces a greater response compared to its single condensate counterpart. Our analysis reveals the functional significance of the condensate size distribution which can be mapped to the recent experimental findings. Given the spatial heterogeneity within condensates and non-linear nature of intracellular networks, we envision MCS to be a generic functional solution, so that structures of network motifs may have evolved to accommodate such configurations.

Bohmian analysis of dark solutions in interfering Bose–Einstein condensates: The dynamical role of underlying velocity fields

In the last decades, the experimental research on Bose–Einstein interferometry has received much attention due to promising technological implications. This has thus motivated the development of numerical simulations aimed at solving the time-dependent Gross–Pitaevskii equation and its reduced one-dimensional version to better understand the development of interference-type features and the subsequent soliton dynamics. In this work, Bohmian mechanics is considered as an additional tool to further explore and analyze the formation and evolution in real time of the soliton arrays that follow the merging of two condensates. An alternative explanation is thus provided in terms of an underlying dynamical velocity field, directly linked to the local phase variations undergone by the condensate along its evolution. Although the reduced one-dimensional model is considered here, it still captures the essence of the phenomenon, rendering a neat picture of the full evolution without diminishing the generality of the description. To better appreciate the subtleties of free versus bound dynamics, two cases are discussed. First, the soliton dynamics exhibited by a coherent superposition of two freely released condensates is studied, discussing the peculiarities of the underlying velocity field and the corresponding flux trajectories in terms of both the peak-to-peak distance between the two initial clouds and the addition of a phase difference between them. In the latter case, an interesting correspondence with the well-known Aharonov-Bohm effect is found. Then, the recurrence dynamics displayed by the more general case of two condensates released from the two opposite turning points of a harmonic trap is considered in terms of the distance between such turning points. In both cases, it is presumed that the initial superposition state is generated by splitting adiabatically a single condensate with the aid of an optical lattice, which is then turned off. Nonetheless, although the lattice does not play any active role in the simulations, the parameters defining the initial states are in compliance with it, which helps in the interpretation and understanding of the results observed.

Microphase Separation Produces Interfacial Environment within Diblock Biomolecular Condensates.

The phase separation of intrinsically disordered proteins is emerging as an important mechanism for cellular organization. However, efforts to connect protein sequences to the physical properties of condensates, i.e., the molecular grammar, are hampered by a lack of effective approaches for probing high-resolution structural details. Using a combination of multiscale simulations and fluorescence lifetime imaging microscopy experiments, we systematically explored a series of systems consisting of diblock elastin-like polypeptides (ELP). The simulations succeeded in reproducing the variation of condensate stability upon amino acid substitution and revealed different microenvironments within a single condensate, which we verified with environmentally sensitive fluorophores. The interspersion of hydrophilic and hydrophobic residues and a lack of secondary structure formation result in an interfacial environment, which explains both the strong correlation between ELP condensate stability and interfacial hydrophobicity scales, as well as the prevalence of protein-water hydrogen bonds. Our study uncovers new mechanisms for condensate stability and organization that may be broadly applicable.

Asymptotic analysis of conversion-limited phase separation

Liquid–liquid phase separation plays a major role in the formation and maintenance of various membrane-less subcellular structures in the cytoplasm and nucleus of cells. Biological condensates contain enhanced concentrations of proteins and RNA, many of which can be continually exchanged with the surrounding medium. Coarsening is an important step in the kinetics of phase separation, whereby an emulsion of polydisperse condensates transitions to a single condensate in thermodynamic equilibrium with a surrounding dilute phase. A key feature of biological phase separation is the co-existence of multiple condensates over significant time scales, which is consistent with experimental observations showing a slowing of coarsening rates. It has recently been proposed that one rate limiting step could be the slow interfacial conversion of a molecular constituent between the dilute and dense phases. In this paper, we analyse conversion-limited phase separation within the framework of diffusion in singularly perturbed domains, which exploits the fact that biological condensates tend to be much smaller than the size of a cell. Using matched asymptotic analysis, we solve the quasi-static diffusion equation for the concentration in the dilute phase, and then derive kinetic equations for the slow growth/shrinkage of the condensates. This provides a systematic way of obtaining corrections to mean-field theory that take into account the geometry of the cell and the locations of all the condensates.

Corrosion Detection and Surface Repair with Coatings on Condensate Storage Tanks Internal Surfaces

In the Nuclear power plant Krško, there are two single hull condensate storage tanks with floating diaphragm each containing up to 757 m3 of demineralized water. The main purpose of these two storage tanks is to provide a capacity of cooling water for cooling the reactor coolant system via steam generators with the use of auxiliary feedwater pumps. This is a very important function from the safety point of view and that is the reason that both storage tanks are listed as safety class 3 components. It is also possible to fill up the condensate system if other means are not available. Condensate storage tanks are subject to periodic testing and periodic inspections to determine the condition of the components. Both tanks are in operation since the start-up of the powerplant, are located outside, and are exposed to different degradation processes. There was a concern that the tanks are leaking because there were often small puddles of water near the tanks. There were no changes in the levels of tanks. The design of tanks is a single hull, so there is no indication if a small leak is present.In the outage 2018, both tanks were emptied and examined with NDE methods to find any corrosion damage of bottom plates and any untight spots on adjacent welds.The article is about the NDE methods that were used (Magnetic Flux Leakage, Ultrasonic and Vacuum Box inspection) to determine the condition of the floor plates and adjacent welds as well as the process of internal surface reparation with coatings. Process of coatings qualification for use in safety class components is also explained: dedication process for material up-grade from non-safety related to safety-related because CY tank linings are classified as safety-related according to RG 1.54, rev 2 and corresponding ASTM standards and NEK technical specification SP-A5001. All activities for surface repair with coatings shall comply with safety-related requirements. Also, extensive immersion tests with selected and specially defined parameters were performed in NEK chemical laboratory in order to select the most suitable coating system for surface repair of CY tank floor lining. Further details concerning immersion tests are presented below.

Biomolecular condensate drives polymerization and bundling of the bacterial tubulin FtsZ to regulate cell division

Cell division is spatiotemporally precisely regulated, but the underlying mechanisms are incompletely understood. In the social bacterium Myxococcus xanthus, the PomX/PomY/PomZ proteins form a single megadalton-sized complex that directly positions and stimulates cytokinetic ring formation by the tubulin homolog FtsZ. Here, we study the structure and mechanism of this complex in vitro and in vivo. We demonstrate that PomY forms liquid-like biomolecular condensates by phase separation, while PomX self-assembles into filaments generating a single large cellular structure. The PomX structure enriches PomY, thereby guaranteeing the formation of precisely one PomY condensate per cell through surface-assisted condensation. In vitro, PomY condensates selectively enrich FtsZ and nucleate GTP-dependent FtsZ polymerization and bundle FtsZ filaments, suggesting a cell division site positioning mechanism in which the single PomY condensate enriches FtsZ to guide FtsZ-ring formation and division. This mechanism shares features with microtubule nucleation by biomolecular condensates in eukaryotes, supporting this mechanism’s ancient origin.

Lifetime of excitations in atomic and molecular Bose-Einstein condensates

Recent experimental progress has produced Molecular Superfluids (MSF) in thermal equilibrium; this opens the door to a new class of experiments investigating the associated thermodynamic and dynamical responses. We review the theoretical picture of the phase diagram and quasiparticle spectrum in the Atomic Superfluid (ASF) and MSF phases. We further compute the parametric dependence of the quasiparticle lifetimes at one-loop order. In the MSF phase, the $U(1)$ particle number symmetry breaks to $\mathbb{Z}_2$ and the spectrum exhibits a gapless Goldstone mode in addition to a gapped $\mathbb{Z}_2$-protected atom-like mode. In the ASF phase, the $U(1)$ symmetry breaks completely, leaving behind a Goldstone mode and an unprotected gapped mode. In both phases, the Goldstone mode decays with a rate given by the celebrated Belyaev result, as in a single component condensate. In the MSF phase, the gapped mode is sharp up to a critical Cherenkov momentum beyond which it emits phonons. In the ASF phase, the gapped mode decays with a constant rate even at small momenta. These decay rates govern the spectral response in microtrap tunneling experiments and lead to sharp features in the transmission spectrum of atoms fired through molecular clouds.

Demixing of distinct ribonucleoprotein condensates is under dynamical control.

Whi3 is an RNA binding protein from Ashbya gossypii that phase separates to form RNA-protein condensates. Observations in cells show that certain RNA molecules co-localize with Whi3 and other RNA molecules into a single condensate, while some others form apparently demixed condensates. Is a there a thermodynamic basis for different ternary combinations of Whi3 and RNA molecules to form demixed condensates? Simulations show that RNA molecules must have strong repulsive interactions for one another, or strong homotypic interactions if demixing is to be thermodynamically feasible. We tested this using in vitro reconstitutions to quantify the phase behaviors of binary and ternary mixtures. A novel analysis of the measured phase boundaries show that heterotypic protein-RNA interactions are the main drivers of phase separation. This was further tested by deletion of cognate binding sites on RNA molecules, showing that there are several ways for Whi3 to associate with RNA molecules. Investigations of phase behaviors internary mixtures show that demixed condensates are dynamically arrested phases. Therefore, in vitro studies suggest that demixing is likely to be under dynamical rather than thermodynamic control. To test for this in live cells, we generated a mutant where differentially expressed RNAs are constitutively expressed under a single promoter. We find that RNA molecules that demix in a wildtype background formed colocalized condensates when they are expressed concomitantly. Our studies show that the non-equilibrium aspects of phase separation, specifically the ability to form dynamically arrested phases, can give rise to demixed condensates that achieve long-livedspatial segregation of RNA molecules within distinct condensates.

Regulation of Polyhomeotic Condensates by Intrinsically Disordered Sequences That Affect Chromatin Binding.

The Polycomb group (PcG) complex PRC1 localizes in the nucleus in condensed structures called Polycomb bodies. The PRC1 subunit Polyhomeotic (Ph) contains an oligomerizing sterile alpha motif (SAM) that is implicated in both PcG body formation and chromatin organization in Drosophila and mammalian cells. A truncated version of Ph containing the SAM (mini-Ph) forms phase-separated condensates with DNA or chromatin in vitro, suggesting that PcG bodies may form through SAM-driven phase separation. In cells, Ph forms multiple small condensates, while mini-Ph typically forms a single large nuclear condensate. We therefore hypothesized that sequences outside of mini-Ph, which are predicted to be intrinsically disordered, are required for proper condensate formation. We identified three distinct low-complexity regions in Ph based on sequence composition. We systematically tested the role of each of these sequences in Ph condensates using live imaging of transfected Drosophila S2 cells. Each sequence uniquely affected Ph SAM-dependent condensate size, number, and morphology, but the most dramatic effects occurred when the central, glutamine-rich intrinsically disordered region (IDR) was removed, which resulted in large Ph condensates. Like mini-Ph condensates, condensates lacking the glutamine-rich IDR excluded chromatin. Chromatin fractionation experiments indicated that the removal of the glutamine-rich IDR reduced chromatin binding and that the removal of either of the other IDRs increased chromatin binding. Our data suggest that all three IDRs, and functional interactions among them, regulate Ph condensate size and number. Our results can be explained by a model in which tight chromatin binding by Ph IDRs antagonizes Ph SAM-driven phase separation. Our observations highlight the complexity of regulation of biological condensates housed in single proteins.

Characterization of RNA content in individual phase-separated coacervate microdroplets

Condensates formed by complex coacervation are hypothesized to have played a crucial part during the origin-of-life. In living cells, condensation organizes biomolecules into a wide range of membraneless compartments. Although RNA is a key component of biological condensates and the central component of the RNA world hypothesis, little is known about what determines RNA accumulation in condensates and to which extend single condensates differ in their RNA composition. To address this, we developed an approach to read the RNA content from single synthetic and protein-based condensates using high-throughput sequencing. We find that certain RNAs efficiently accumulate in condensates. These RNAs are strongly enriched in sequence motifs which show high sequence similarity to short interspersed elements (SINEs). We observe similar results for protein-derived condensates, demonstrating applicability across different in vitro reconstituted membraneless organelles. Thus, our results provide a new inroad to explore the RNA content of phase-separated droplets at single condensate resolution.

Holographic multicondensate with nonlinear terms

We study the influence of nonlinear terms quartic of the condensates on the phase structure of a holographic model with a multicondensate in the probe limit. We include one s-wave order and one p-wave order charged under the same U(1) gauge field in the holographic model and study the influence of the three quartic nonlinear terms with coefficients ${\ensuremath{\lambda}}_{s}$, ${\ensuremath{\lambda}}_{p}$ and ${\ensuremath{\lambda}}_{sp}$ on the phase structure. The quartic terms in action result in cubic nonlinear terms in equations of motion, which do not affect the critical points of a single condensate because the infinitesimal condensate value vanishes the nonlinear contributions. However, the quartic terms have a clear influence on the phase structure of systems containing multicondensates. We show the influence of each of the three parameters on the phase diagram with the other two set to zero, respectively. With these nonlinear terms, we get new power on tuning the phase structure of the holographic systems showing multicondensates, and realize a reentrant phase transition as an example.

Compartmentalization of telomeres through DNA-scaffolded phase separation.

SummaryTelomeres form unique nuclear compartments that prevent degradation and fusion of chromosome ends by recruiting shelterin proteins and regulating access of DNA damage repair factors. To understand how these dynamic components protect chromosome ends, we combine in vivo biophysical interrogation and in vitro reconstitution of human shelterin. We show that shelterin components form multicomponent liquid condensates with selective biomolecular partitioning on telomeric DNA. Tethering and anomalous diffusion prevent multiple telomeres from coalescing into a single condensate in mammalian cells. However, telomeres coalesce when brought into contact via an optogenetic approach. TRF1 and TRF2 subunits of shelterin drive phase separation, and their N-terminal domains specify interactions with telomeric DNA in vitro. Telomeric condensates selectively recruit telomere-associated factors and regulate access of DNA damage repair factors. We propose that shelterin mediates phase separation of telomeric chromatin, which underlies the dynamic yet persistent nature of the end-protection mechanism.

Semi-analytical model for the heat conduction resistance of a single spherical condensate droplet

The evolution and heat transfer of a single spherical condensate droplet on a superhydrophobic surface (SHS) can be modeled by the droplet thermal resistance network. The calculation of the heat conduction resistance of a single spherical condensate droplet (Rd) is a fundamental problem in analyzing the droplet thermal resistance network. Till now, the majority of researchers have used Kim's formula to calculate Rd. However, the results from Kim's formula have errors above 85% compared to those from the CFD software (ANSYS Fluent). In this paper, we analyze the reason for inaccuracy of Kim's formula. Taking two-dimensional droplet heat conduction into consideration, we gain the theoretical temperature field function inside the condensate droplet by utilizing the conformal mapping. Basing on calculation of heat transfer rate through the droplet and ANSYS Fluent analysis, we put forward a new semi-analytical model to calculate Rd. We verify and find that the results from our new model have errors less than 13% compared to those from ANSYS Fluent. It means that our new semi-analytical model is more accurate than Kim's formula. We calculate the Rd proportion of the overall heat transfer resistance and find that the application of our model can obviously decrease heat transfer calculation errors for big droplets (r>16μm) with large contact angles (θ>120∘). The new semi-analytical model for the heat conduction resistance of a single spherical condensate droplet is promising in the sense that it might replace Kim's model to become a novel design criterion of enhanced heat transfer SHSs.

Observation of fragmentation of a spinor Bose-Einstein condensate.

Weakly interacting Bose gases usually form Bose-Einstein condensates in which most particles occupy the same single-particle state. However, when this state cannot realize a continuous symmetry of the many-body Hamiltonian, a fragmented condensate exhibiting the expected symmetry may emerge. Here, we produced a three-fragment condensate for a mesoscopic spin-1 gas of about 100 atoms, with anti-ferromagnetic interactions and vanishing collective spin. Using a spin-resolved detection approaching single-atom resolution, we show that the reconstructed state is close to the expected many-body ground state, whereas one-body observables are the same as for a completely mixed state. Our results highlight how the interplay between symmetry and interactions generates entanglement in a mesoscopic quantum system.

Covalent Conjugation of Carbon Dots with Plasmid and DNA Condensation Thereafter: Realistic Insights into the Condensate Morphology, Energetics, and Photophysics.

The use of carbon quantum dots (CDs) as trackable nanocarriers for plasmid and gene as hybrid DNA condensates has gained momentum, as evident from the significant recent research efforts. However, the in-depth morphology of the condensates, the energetics of the condensation process, and the photophysical aspects of the CD are not well understood and often disregarded. Herein, for the first time, we covalently attached linearized pUC19 with citric acid and cysteamine-derived CD through the reaction of the surface amine groups of CDs with the 5′-phospho-methyl imidazolide derivative of the plasmid to obtain a 1:1 CD-pUC19 covalent conjugate. The CD-pUC19 conjugates were further transformed into DNA condensates with spermine that displayed a toroidal morphology with a diameter of ∼200 nm involving ∼2–5 CD-pUC19 conjugates in a single condensate. While the interaction of pristine CD to spermine was exothermic, the binding of the CD-pUC19 conjugate with spermine was endothermic and primarily entropy-driven. The condensed plasmid displayed severe conformational stress and deviation from the B-form due to the compact packing of the DNA but better transfection ability than the pristine CD. The CDs in the condensates tend to come close to each other at the core that results in their shielding from excitation. However, this does not prevent them from emanating reactive oxygen species on visible light exposure that compromises the decondensation process and cell viability at higher exposure times, calling for utmost caution in establishing them as nonviral transfecting agents universally.

Influence of different categories of condensates on film condensation of R-134a on in-line arranged 3D finned tubes

The condensates generated in the process of film condensation, such as condensing, falling, and acting condensates, are identified and defined to obtain precise inundation effect model of 3D finned tube bundle. Furthermore, the influence of three categories of condensates on the film condensation under different heat fluxes and condensing temperatures is experimentally investigated via the homologous method. A test bench with a test section contains two arrays of 8 horizontal tubes is initially constructed. Experimental results show that the inundation effect coefficient decreases from 1.00 to 0.78 as acting condensate Re increases from 75 to 1860. The influence of heat flux on three categories of condensates is more significant than that of condensing temperature. As tube row depth increases from 1 to 6, the proportion of condensing condensate Re to acting condensate Re decreases from 100% to 15.0%, whereas the proportion of falling condensate Re to acting condensate Re increases from 0 to 85.0%. Furthermore, the condensation heat transfer model of single 3D finned tube and inundation effect model are established, and the deviations of results between experiments and models are almost within ±3.0%. The deviations of inundation effect coefficients between experiment and model proposed in this study are 92.5% and 50.0% less than those between experiment and models using routine method and considering single condensate with homologous method, respectively. Moreover, the inundation effect model proposed in this study reveals that different proportions of condensing or falling condensates to acting condensate lead to different inundation effect coefficient at the same acting condensate Re. The research results will lay the foundation for further revealing the mechanism of film condensation on 3D finned tube bundle and guide the thermal design of 3D finned tubes applied in shell-and-tube condensers.

Plane- and stripe-wave phases of a spin-orbit-coupled Bose-Einstein condensate in an optical lattice with a Zeeman field

A weakly interacting, spin-orbit coupled, ultracold, dilute Bose gas on a two-dimensional square lattice with an external Zeeman field is studied. We explore the plane and stripe wave phases of the system involving nonzero condensate momenta, which occur when the Zeeman field is below a critical value. Their excitation spectra are found using Bogoliubov theory and by two different routes. The validity of each method to obtain the excitation spectrum is discussed, and it is found that projection on the lowest single-particle band is an excellent approximation in the plane wave phase, while it is a poor approximation in the stripe wave phase. While the plane wave phase has a phonon minimum at its single condensate momentum, revealing a nonzero sound velocity of the excitations, the stripe wave phase has quadratic minima at its two condensate momenta showing zero sound velocity of the excitations. We discuss how the presence of more than one condensate momentum is essential for these differences between the two phases. Additionally, it is emphasized that the zero sound velocity in the stripe wave phase is a lattice effect, since continuum studies of the same phase have shown nonzero sound velocity.

Differential interferometry using a Bose-Einstein condensate

Out of a single Bose-Einstein condensate (BEC), we create two simultaneous interferometers, as employed for the differentiation between rotations and accelerations. Our method exploits the precise motion control of BECs combined with the precise momentum transfer by double Bragg diffraction for interferometry. In this way, the scheme avoids the complexity of two BEC sources and can be readily extended to a six-axis quantum inertial measurement unit.Graphical abstract

The Douglas Field, Block 110/13b, UK East Irish Sea

Abstract The Douglas Field is located in Block 110/13b in the East Irish Sea. It was the first oilfield to be discovered and produced in the region, having been found in 1990 and brought on stream in January 1996. The field structure comprises a series of north–south-trending, tilted, extensional fault blocks. The reservoir interval is the Triassic Ormskirk Sandstone Formation comprising good quality aeolian and fluvial sandstones. The field is relatively shallow, with the top reservoir at c. 2120 ft true vertical depth subsea. The hydrocarbon is a light oil of 44°API gravity with a maximum column height of c. 400 ft. The Douglas Field contains an estimated stock tank oil in place of 248 MMbbl and was developed with 22 wells: 15 producers, six water injectors and a single sour gas and condensate disposal well. Electric submersible pumps are installed in oil producers for artificial lift and water injection was utilized from field start-up for pressure maintenance. A water-alternating-gas pilot was implemented on the field in 2017 as an enhanced oil recovery scheme. The field currently produces at a rate of c. 4000 bopd, with approximately 90% water cut. The field has produced 103 MMbbl to date, giving a current oil recovery of c. 41%.