Macromolecular Components Research Articles

Evaluation of the Cultivated Mushroom Pleurotus ostreatus Basidiocarps Using Vibration Spectroscopy and Chemometrics

Fruiting bodies (basidiocarps) of the cultivated mushroom Pleurotus ostreatus (16 strains) were characterized by vibration spectroscopy and chemometrics. According to organic elemental analysis and Megazyme assay, the basidiocarps contained ~6.2–17.5% protein and ~18.8–58.2% total glucans. The neutral sugar analysis confirmed that glucose predominated in all the samples (~71.3–94.4 mol%). Fourier-transformed (FT) mid- and near-infrared (FT MIR, FT NIR) and FT Raman spectra of the basidiocarps were recorded, and the characteristic bands of proteins, glucans and chitin were assigned. The samples were discriminated based on principal component analysis (PCA) of the spectroscopic data in terms of biopolymeric composition. The partial least squares regression (PLSR) models based on first derivatives of the vibration spectra were obtained for the prediction of the macromolecular components, and the regression coefficients R2 and root mean square errors (RMSE) were calculated for the calibration (cal) of proteins (R2cal 0.981–0.994, RMSEcal ~0.3–0.5) and total glucans (R2cal 0.908–0.996, RMSEcal ~0.6–3.0). According to cross-validation (CV) diagnosis, the protein models were more precise and accurate (R2cv 0.901–0.970, RMSEcv ~0.6–1.1) than the corresponding total glucan models (R2cv 0.370–0.804, RMSEcv ~4.7–8.5) because of the wide structural diversity of these polysaccharides. Otherwise, the Raman band of phenylalanine ring breathing vibration at 1004 cm−1 was used for direct quantification of proteins in P. ostreatus basidiocarps (R ~0.953). This study showed that the combination of vibration spectroscopy with chemometrics is a powerful tool for the evaluation of culinary and medicinal mushrooms, and this approach can be proposed as an alternative to common analytical methods.

Effect of mechanical homogenization on the physicochemical properties of films made from dual modified corn starch prepared by the casting solution method

Films were obtained by casting a solution of succinated hydrolyzed corn starch homogenized using an ULTRA-TURRAX (UF). Glycerol was added as a plasticizer. Films obtained after homogenizing with a magnetic bar by the conventional method (CF) were used as a control. The physicochemical, thermomechanical, and hygroscopic properties were evaluated. UF resulted in a gray color change when heating at 65°C during the mechanical homogenization process, while the CF remained white. The thermomechanical properties depended on the conditioning at different relative humidity values. Mechanical properties of the UF showed the lowest values of tensile strength (TS) and percentage of elongation (%E). The mechanical homogenization at high revolutions allowed changing the physicochemical and mechanical properties of the films obtained from dual modified starch, probably due to the dispersion, rearrangement, orientation, and interaction of the macromolecular components of the modified polymers. These films could have a wide array of applications. Practical applications These films have a wide array of applications in foods with low moisture content. Also providing new information for future research using doubly modified starch by homogenizing it at high revolutions, causing new molecular interactions and eventually physicochemical properties new.

Skin hydration dynamics investigated by electrical impedance techniques in vivo and in vitro

Skin is easily accessible for transdermal drug delivery and also attractive for biomarker sampling. These applications are strongly influenced by hydration where elevated hydration generally leads to increased skin permeability. Thus, favorable transdermal delivery and extraction conditions can be easily obtained by exploiting elevated skin hydration. Here, we provide a detailed in vivo and in vitro investigation of the skin hydration dynamics using three techniques based on electrical impedance spectroscopy. Good correlation between in vivo and in vitro results is demonstrated, which implies that simple but realistic in vitro models can be used for further studies related to skin hydration (e.g., cosmetic testing). Importantly, the results show that hydration proceeds in two stages. Firstly, hydration between 5 and 10 min results in a drastic skin impedance change, which is interpreted as filling of superficial voids in skin with conducting electrolyte solution. Secondly, a subtle impedance change is observed over time, which is interpreted as leveling of the water gradient across skin leading to structural relaxation/changes of the macromolecular skin barrier components. With respect to transdermal drug delivery and extraction of biomarkers; 1 h of hydration is suggested to result in beneficial and stable conditions in terms of high skin permeability and extraction efficiency.

Pectin biosynthesis pathways are adapted to higher rhamnogalacturonan formation in lignocellulosic jute (Corchorus spp.)

Pectin and lignin are two enigmatic macromolecular components of cell wall, which are spatially and temporally deposited during plant growth. While the former is important for primary growth, the latter accumulates during secondary growth. With the evolution of land plants, the structural complexity of pectin increased to protect cell walls against the intrusion of pectinolytic pathogens besides offering multiple choices for its interaction with lignin. We reconstructed pectin biosynthesis pathways in an annual lignocellulosic bast fibre-producing crop jute (Corchorus spp.) from hypocotyl transcriptomes and identified 27 isoforms of 17 genes and 12 isoforms of galacturonosyltransferase involved in nucleotide-sugar interconversion and pectin polymerization, respectively. Salvage pathways were found to be functional for replenishing nucleotide sugars in jute hypocotyls. Phylogenetic analyses revealed that the genes of pectin biosynthesis pathways are well conserved across taxa. For significant upregulation of the rhamnose biosynthesis gene (RHM1), we traced its evolution, identified its two-domain protein structure proposed to have been evolved from charophycean green algae and generated a structural model explaining its functional conservation. Retting of jute stem with pectinolytic bacteria showed that pectin complexity and its interaction with lignin play crucial roles in resisting cell wall deconstruction that may have accrued from increased rhamnogalacturonan biosynthesis.

Membrane fouling control by UV/persulfate in tertiary wastewater treatment with ultrafiltration: A comparison with UV/hydroperoxide and role of free radicals

The membrane fouling caused by effluent organic matter (EfOM) is a bottleneck for application of ultrafiltration (UF) in tertiary wastewater treatment. In this study, two ultraviolet-based oxidation pretreatments, i.e., ultraviolet/persulfate (UV/PS) and ultraviolet/hydrogen peroxide (UV/H2O2), were used to alleviate EfOM fouling at a low UV dose of 2.36 × 103 mJ cm−2, in consideration of the widely use of UV radiation for disinfection in wastewater treatment plants. Effects of the pretreatments on characteristics of EfOM were investigated in terms of fluorescence spectrum, molecular weight distribution, and hydrophobicity. Moreover, filtration tests were performed to assess impacts of the oxidation pretreatments on flux decline, fouling reversibility, and fouling mechanisms. The results showed that the UV/PS pretreatment substantially reduced flux decline and improved fouling reversibility during filtration of the secondary effluent, attributed to efficient degradation (~67%, in terms of UV254) and partial mineralization (~39%, in terms of DOC) of EfOM. In particular, the irreversible resistance was decreased by nearly one order of magnitude at the PS dose of 1.0 mM. UV/H2O2 was not effective in EfOM fouling control under the conditions investigated. The UV/PS pretreatment resulted in a shift of mechanisms for EfOM fouling from successive pore blocking and cake filtration to single standard blocking, attributing to preferential degradation of macromolecular components in EfOM and a weakened role of cake interception. The radical quenching experiment showed that the sulfate radicals played a dominant role in EfOM fouling control by UV/PS, and that transformation of sulfate radicals into hydroxyl radicals negatively affected EfOM fouling control.

Culture of pig embryos

Pig embryos can be cultured using a number of different strategies including complex approaches like culture in vivo in a surrogate oviduct (rabbit, sheep, mouse), culture in mouse oviducts in organ culture, and co-culture of embryos with cells in addition to simple approaches like culture in defined media or salt solutions. Addition of serum to medium has been of particular importance where blastocyst development and hatching are required. Pig conceptuses (day 10-15), embryonic discs or cell lines derived from conceptuses can be cultured in complex media like Eagle's minimal essential medium or Dulbecco's modified Eagle's medium with serum, although embryonic discs can be cultured in the absence of serum. In contrast, early stage pig embryos (one-cell to blastocyst) are best cultured in simpler media such as those used for mouse embryos. The media that have been used are all relatively similar in composition. They contain salts and one or more energy sources such as glucose, lactate, or pyruvate with BSA as a macromolecular component. Early attempts to culture pig embryos were not very successful, but some embryos did develop to the blastocyst stage. More recent reports indicate a much higher rate of development with relatively little or no change in media composition. Some workers have reported improved development in medium lacking glucose, which is consistent with findings with laboratory animals such as hamsters. Glutamine can serve as the sole exogenous energy source in medium lacking glucose, lactate and pyruvate. Addition of taurine and hypotaurine to culture medium enhances development of pig embryos in vitro. We suggest, where possible, adoption of a standard medium that could be used by different laboratories and, perhaps, with different species. Use of one medium for different species would simplify experimental protocols, enhance studies of comparative embryonic physiology and metabolism, and expedite transfer of information obtained in different species.

Reactive oxygen species are required for spore wall formation in Physcomitrella patens.

A robust spore wall was a key requirement of terrestrialization by early plants. Sporopollenin in spore and pollen grain walls is thought to be polymerized and cross-linked to other macromolecular components partly through oxidative processes involving H2O2. Therefore, we investigated effects of scavengers of reactive oxygen species (ROS) on formation of spore walls in the moss, Physcomitrella patens. Exposure of sporophytes, containing spores in the process of forming walls, to ascorbate, dimethylthiourea or 4-hydroxy-TEMPO prevented normal wall development in a dose, chemical and stage-dependent manner. Mature spores, exposed while developing to a ROS scavenger, burst when mounted in water on a flat slide under a coverslip (a phenomenon we named "augmented osmolysis" since they did not burst in phosphate-buffered saline or in water on a depression slide). Additionally, walls of exposed spores were more susceptible to alkaline hydrolysis than those of control spores and some were characterized by discontinuities in the exine, anomalies in perine spine structure, abnormal intine and aperture and occasionally wall shedding. Our data support involvement of oxidative cross-linking in spore wall development, including sporopollenin polymerization or deposition, as well as a role for ROS in intine/aperture development.

TG- MS study on in-situ sulfur retention during the co-combustion of reclaimed asphalt binder and wood sawdust

Reclaimed asphalt binder (RAB) releases large amounts ·of hazardous sulfur-containing gases during combustion. This study attempts to introduce wood sawdust (WS) as an in-situ inhibitor of sulfur release during the combustion of refuse-derived fuel (RDF) blended with RAB-WS. The combustion characteristics, gaseous sulfur-containing products, interactions and combustion kinetics of RDF were investigated through thermogravimetry and mass spectrometry (TG-MS), and the mechanisms on migration and distribution of sulfur were revealed. Results indicated that WS additive inhibits the volatilization of light components and promotes the degradation of macromolecular components. WS addition improved the combustibility, burnout performance and combustion stability of RAB. The sulfur release of RAB-based RDF was mainly derived from resins and asphaltenes. WS addition generally decreased all gaseous sulfur-containing compounds (CH3SH, COS, SO2, CS2 and thiophene). Interactions between RAB and WS restrained all sulfur-containing gas emissions, and the normalized sulfur inhibition ratio reached 40.99 %. The Sarink and DAEM models could well describe the kinetic process of the co-combustion of RAB and WS. WS addition led to a decrease in activation energy, namely, it lowered the reaction barrier. Sulfur could be retained in-situ into incineration residue through the formation of sulfate minerals during the co-combustion of RAB and WS.

Elevated MUC1 and MUC5AC mucin protein levels in airway mucus of critical ill COVID-19 patients.

Coronavirus disease 2019 (COVID-19) patients exhibit a spectrum of respiratory symptoms like cough and dyspnea1-3. Airway mucus is an adhesive viscoelastic gel composed mostly of high-molecular weight mucous glycoproteins and water, which is important in maintaining lung function and health, pathological mucus hypersecretion may cause airway obstruction and lead to respiratory distress. Mucin (MUC) glycoproteins are the major macromolecular components of mucus, which are classified into two major types: the gel-forming, secreted mucins (i.e, MUC5AC) and the membrane-tethered mucins (i.e. MUC1)4. Here, with an attempt to understand the lung changes, we sought to provide a delineation of the components of airway mucus from COVID-19 patients. This article is protected by copyright. All rights reserved.

Fractionation and characterization of polyphenolic compounds and macromolecules in red wine by asymmetrical flow field-flow fractionation.

Red wine is a complex matrix containing macromolecules such as condensed tannins and polysaccharides. Wine macromolecular components and their interactions have been reported to impact taste properties such as astringency but the colloidal systems formed in wine are not well known. A key prerequisite to characterize these systems is the ability to work under analytical conditions as close as possible to the colloid environment, preserving the sample structure and limiting the denaturation of macromolecular complexes. A method of Asymmetric Flow Field-Flow Fractionation (AF4) coupled with UV detection, multi-angle light scattering (MALS), and differential refractometer index (dRI) (AF4-UV-MALS-dRI) has been developed to analyse macromolecules, including tannins and polysaccharides, and macromolecular complexes, in red wine. This method separates objects according to their hydrodynamic radius and does not require calibration to determine molecular weight (Mw). AF4 can provide native separation of wine colloidal matter while working with simulated wine as mobile phase. The channel was equipped with a 350-µm spacer and the membrane made in regenerated cellulose had a cut-off of 5kDa. Different parameters of crossflow rate were investigated using a generic red wine to optimize separation conditions. Then, purified fractions of polysaccharides and tannins were analysed using the selected AF4 parameters. The comparison of the peaks obtained for these fractions and for the wine sample allowed us to determine the retention time associated with these macromolecules. The AF4 fractogram of wine was divided into four fractions. The first three were assigned to higher Mw tannins coeluted with lower Mw polysaccharides such as rhamnogalacturonan II (F1), to intermediate Mw polysaccharides (F2), and to higher Mw mannoproteins (F3) whereas the last fraction (F4) was not identified. Furthermore, our results have shown that AF4-UV-MALS-dRI could be an efficient technique to separate large size tannins as well as polysaccharides and macromolecular complexes.

Understanding the torrefaction of woody and agricultural biomasses through their extracted macromolecular components. Part 2: Torrefaction model

A new torrefaction model was proposed for predicting solid mass loss in torrefaction as a function of biomass main macromolecular composition and type, as well as on the operating conditions. To do this, solid degradation kinetics were modelled following a 2-successive reaction scheme for each macro-compound and the additive modelling approach through biomass macromolecular component behavior in torrefaction proposed by Nocquet et al. (2014). The use of extracted fractions from different woody and agricultural biomass species (ash-wood, beech, miscanthus, pine and wheat straw) instead of commercial compounds increased the accuracy of the prediction of solid kinetics in biomass torrefaction. The validation of the proposed model with 9 raw biomasses in torrefaction showed an accurate prediction for woods, while the prediction for agricultural biomasses was acceptable.

Screening and structure study of active components of Astragalus polysaccharide for injection based on different molecular weights

As a special traditional Chinese medicine, Astragalus polysaccharides for injection (APS, batch no. Zhunzi Z20040086) includes complex polysaccharide macromolecules that may increase the risk upon application. Although fingerprints for quality control are available, the specific active ingredients are unclear. Identifying the active components is the key to reduce the risk of adverse reactions of the drug. In this work, APS was mainly separated into two components, namely, macromolecular component (APS-I) and small molecular components (APS-II). The molecular weight measurement revealed that the average molecular weight of APS-I exceeded 500 kDa, and that of APS-II was 10 kDa. Monosaccharide-composition analysis revealed that APS-I consisted of glucose, galactose, arabinose, rhamnose, and galacturonic acid, with a ratio of approximately 1.5:1:5.4:0.08:0.1. Meanwhile, APS-II consisted of glucose, galactose, arabinose, rhamnose, and galacturonic acid, with a molar ratio of 9:1:1.4:0.04:0.001. Methylation, FT-IR, and NMR analysis indicated that the APS-I monosaccharide residue was linked as follows: D-Glcp-(1→, →4)-D-Glcp-(1→, →2)-L-Rhap-(1→, D-Araf-(1→, →5)-D-Araf-(1→, →2,5)-D-Araf-(1→, →4)-D-Galp-(1 → . Meanwhile, the APS-II monosaccharide residue was connected as follows: α-D-Glcp-(1→, →4)-α-D-Glcp-(1→, →6)-α-D-Glcp-(1→, →4,6)-α-D-Glcp-(1→, →3,4,6)-α-D-Glcp-(1→, →2)-α-L-Rhap-(1→, α-D-Araf-(1→, →5)-α-D-Araf-(1→, →4)-β-D-Galp-(1 → . Screening experiments on their in vitro immunological activity showed that APS-II had stronger effect on innate and adaptive immunities than APS-I. In vivo animal experiments showed that APS-II can increase the leukocyte level of cyclophosphamide immunosuppressed mice and improve their immunomodulatory ability. Therefore, APS-II is the main active ingredient of APS and is expected to become a new generation of APS products.

Understanding the torrefaction of woody and agricultural biomasses through their extracted macromolecular components. Part 1: Experimental thermogravimetric solid mass loss

The behavior of biomass in torrefaction is determined by that of its macromolecular components, as well as by the biomass type. However, up to now, commercial compounds were typically used for modelling biomass torrefaction. This work proposes to assess the behavior of cellulose, hemicelluloses and lignin in torrefaction through extracted fractions directly isolated from woody and agricultural biomasses (ash-wood, beech, miscanthus, pine and wheat straw) thanks to an optimized extraction procedure. The solid kinetics of these extracted fractions were analyzed through thermogravimetric analysis (TGA) in chemical regime conditions (200–300 °C at 3 °C·min−1 followed by 30 min at 300 °C). These experiments highlighted the influence of the biomass type and the sugar composition in the degradation of the polysaccharide fractions in torrefaction, particularly for hemicelluloses. Furthermore, the degree of preservation of the native structure of the macromolecular components, when extracting them from biomass, seems also having an impact their behavior, especially for cellulose. The comparison of the torrefaction solid kinetic profiles of these extracted fractions, dependent on the biomass type, to that of commercial compounds from previous studies suggest that these extracted fractions would be more suitable for biomass torrefaction modelling.

Evolution of Two Ubiquitin-like System of Autophagy in Orchid

As an important horticultural plant, the orchid is widely distributed in its natural habitat and faces various environmental stresses, among which nutrient recycling and stress resistance are of great concern. During these processes, autophagy is an essential pathway, which is a conserved self-eating process that degrades macromolecular components and recycles cell materials or nutrients during developmental processes or under stress conditions. Two ubiquitin-like systems (UBLs) play a major role in the initiation of autophagy and are associated with two key proteins: ATG8 and ATG12. In this study, we identified and refined the UBL-related genes in orchids and performed phylogenetic reconstruction together with other plant species. We found that the orchid had unique domains in UBL-related genes, indicating potential functional diversification in the ATG8 system in plants. Transcriptome and protein tertiary structure prediction indicated that conserved domains that are vital for the canonical function of ATG12 are incomplete in orchids, in which a novel mechanism of autophagy may have evolved.

Functional DNA Structures and Their Biomedical Applications

Since the discovery of the double-helix structure in 1953, nucleic acids have been developed from natural genetic codes into functional building blocks in a wide range of biotechnology and material...

Several Sterilization Strategies Maintain the Functionality of Mucin Glycoproteins.

Mucin glycoproteins, the macromolecular components of mucus, combine a broad range of biomedically important properties. Among those is the ability of mucin solutions to act as excellent lubricants. However, to be able to use purified, endogenous mucin glycoproteins as components of a biomedical product, the mucins need to be sterile; this, in turn, makes it necessary to subject the mucins to quite harsh physical treatments, such as heat exposure, autoclaving, UV-, or γ-irradiation, which might compromise the functionality of the glycoproteins. Here, it is shown that mucins are indeed able to withstand most of those treatments without suffering significant lubrication impairment or structural degradation. Among those treatments, which left the mucins unharmed, γ-irradiation is identified to be the most powerful one in terms of inactivating microbial contaminations. The obtained results demonstrate a remarkable sturdiness of mucins, which opens up broad possibilities for them to be further processed into materials, e.g., as parts of biomedical products.

Muc5ac Expression Protects the Colonic Barrier in Experimental Colitis.

The mucus gel layer (MGL) lining the colon is integral to exclusion of bacteria and maintaining intestinal homeostasis in health and disease. Some MGL defects allowing bacteria to directly contact the colonic surface are commonly observed in ulcerative colitis (UC). The major macromolecular component of the colonic MGL is the secreted gel-forming mucin MUC2, whose expression is essential for homeostasis in health. In UC, another gel-forming mucin, MUC5AC, is induced. In mice, Muc5ac is protective during intestinal helminth infection. Here we tested the expression and functional role of MUC5AC/Muc5ac in UC biopsies and murine colitis. We measured MUC5AC/Muc5ac expression in UC biopsies and in dextran sulfate sodium (DSS) colitis. We performed DSS colitis in mice deficient in Muc5ac (Muc5ac-/-) to model the potential functional role of Muc5ac in colitis. To assess MGL integrity, we quantified bacterial-epithelial interaction and translocation to mesenteric lymph nodes. Antibiotic treatment and 16S rRNA gene sequencing were performed to directly investigate the role of bacteria in murine colitis. Colonic MUC5AC/Muc5ac mRNA expression increased significantly in active UC and murine colitis. Muc5ac-/- mice experienced worsened injury and inflammation in DSS colitis compared with control mice. This result was associated with increased bacterial-epithelial contact and translocation to the mesenteric lymph nodes. However, no change in microbial abundance or community composition was noted. Antibiotic treatment normalized colitis severity in Muc5ac-/- mice to that of antibiotic-treated control mice. MUC5AC/Muc5ac induction in the acutely inflamed colon controls injury by reducing bacterial breach of the MGL.

Mechano-Vibrational Spectroscopy of Tissues and Materials Using Vibrational Optical Coherence Tomography: A New Non-Invasive and Non-Destructive Technique

Elastic moduli of tissues and synthetic polymeric materials are important design properties needed to develop new implants. In this paper we report the use of vibrational optical coherence tomography (VOCT) to measure the resonant frequency and calculate the modulus of tissues and synthetic polymers non-invasively and non-destructively in vitro and in vivo. Values of tissue and polymer moduli were obtained by applying an audible sinusoidal sound wave to the surface of each specimen. The sound wave travels to the interior of the material and is reflected back to the surface from underlying layers. A spectrum of resonant frequencies and moduli are then obtained. By analyzing the frequencies at which the maximum displacements are observed the resonant frequencies are determined of the material components and the moduli can then be calculated from calibration equations. The characteristic spectrum of resonant frequencies measured is a finger print by which the composition of the underlying tissue components can be determined using VOCT. The resonant frequencies of cells and tissues range from about 40 Hz for fat to about 990 Hz for lamellar bone. The modulus for these tissues ranges from about 1 MPa (cells) to that for extracellular matrix components that range from 2 to 173 MPa (tissue macromolecular components). Measurements reported on synthetic polymers indicate that the resonant frequency ranges from 80 Hz (silicone rubber) to 2800 Hz (acrylonitrile butadiene styrene) and values of the modulus are found to be between 2 MPa and 2120 MPa, respectively. The resonant frequency and modulus are shown to decrease as a result of fatigue in Viton rubber. It is concluded that the width of the resonant frequency peak is related to the molecular weight distribution and that VOCT may serve as method to rapidly determine molecular weight distributions and properties after cyclic loading of uncrosslinked and crosslinked polymers.

Aging behaviors of bitumen degraded by the microbial consortium on bituminous pavement

As bituminous pavement is under the shady, humid or polluted environments, the bitumen undergoes the microbial degradation by different microorganisms to cause the aging. To understand the microbial aging of bitumen degraded by multiple microorganisms, the typical bacillus licheniformis (BL), bacillus subtilis (BS) and pseudomonas aeruginosa (PA) were selected to compound three microbial consortia in different proportions, and cultured in three culture mediums. A more suitable culture medium for the three microbial consortia is selected. The changes in microscopic morphology, components, molecular weight distributions, functional groups and micromechanical properties of aged bitumen were discussed. Results indicate that the peptone content change of culture medium obviously affects the propagation of microbial consortia. The prepared culture medium II meets the propagation requirements of the three microorganisms, improving their degradation ability. Additionally, the microbial consortia preferentially degrade bituminous light components as their nutrient sources, and show weak degradation ability to macromolecular components, producing potholes with layer structures on bituminous surface. Moreover, the smaller molecules are decreased, but the macromolecules are increased in aged bitumen. The molecular weight distribution of aged bitumen is narrowed. BS, BL and PA in the microbial consortium of S2 show better synergistic effect to efficiently degrade bitumen than those of S1 and S3. Finally, the carbonyl and sulfoxide contents are not changed obviously, but the methylene content is decreased to reflect the aging results of bitumen. The maximum adhesive force of aged bitumen is lowered to cause the decrease in adhesion work, reflecting the decrease in bituminous viscidity. The elastic modulus of bitumen shows an increasing trend as the aging duration is prolonged. The elastic modulus of aged bitumen by the microbial consortium of S2 is higher than those by S1 and S3. This study provides an insight into the microbial aging mechanism of bitumen by multiple microorganisms.

Tug of War between Condensate Phases in a Minimal Macromolecular System.

Membraneless organelles, comprising dozens to hundreds of macromolecular components, form heterogeneous phases in space and evolve over time in material properties. Here, using four macromolecules, we demonstrate a range of phase behaviors associated with membraneless organelles and uncover the underlying physicochemical rules. The macromolecules are SH35 (S) and PRM5 (P), two pentameric, oppositely charged protein constructs; heparin (H), an anionic polymer; and lysozyme (L), a cationic single-domain protein. The S:P, S:L, and P:H binaries form droplets, but the H:L binary forms network-like precipitates, therefore setting up a tug of war between different condensate phases within the S:P:H:L quaternary. The H:L exception can partly be attributed to the compactness of L, as supported by ThT binding data. Increasing amounts of P alone or both S and P, but not S alone, can dissolve H:L precipitates into droplets. These differential effects can be explained by the order of the strengths of pairwise attraction: H:L > P:H > S:P > S:L, deduced from the shapes of ternary phase boundaries. When S and P are at subdissolution concentrations, S:P:H:L precipitates change over time to become droplet-like in appearance, although not completely fluidic according to fluorescence recovery after photobleaching. In fact, confocal microscopy reveals separated S:L-rich and P:H-rich foci inside the droplet-like condensates. Therefore, complex phase behaviors of membraneless organelles, including rescue of aberrant phase transitions, demixing of condensates, and time evolution of material properties, can all be reconstituted and understood via a minimal macromolecular system.