Liquid condensate is a common state of proteins and polypeptides at the regime of high intermolecular interactions

Liquid-liquid phase separation (LLPS) has emerged as a crucial biological mechanism for sequestering macromolecules (such as proteins and nucleic acids) into membraneless organelles in cells. Unstructured and intrinsically disordered domains are known to facilitate multivalent interactions driving protein LLPS. We hypothesized that LLPS could be an intrinsic property of proteins/polypeptides at their high intermolecular interaction regime. To examine this, we studied many (a total of 23) proteins/polypeptides with different structures and sequences for LLPS study using molecular crowder polyethylene glycol (PEG-8000). We showed that all proteins and even highly charged polypeptides (under study) can undergo liquid condensate formation, however with different phase space and conditions. Using a single component and combinations of protein multicomponent (co-LLPS) systems, we establish that a variety of intermolecular interactions can drive proteins/polypeptides LLPS.

Thiophene extraction in PEG - hexan - water based systems

The effect of individual and multicomponent additives of sodium chloride, iron chloride, sodium nitrate and 1-methyl-2-pyrrolidone on the interfacial distribution of thiophene in the systems polyethylene glycol 1500 (PEG-1500) - hexane - water and polyethylene glycol 8000 (PEG-8000) - hexane - water has been studied in this investigation. It was found that in most cases, the introduction of a small amount of individual and joint additives increases the extraction of thiophene from the organic phase to the aqueous.

Investigation of the Influence of the Molecular Weight of Polyethyleneglycols on the Optical Properties and Dispersed Characteristics of Sols of Au Nanoparticles used in Medicine

In this work, the synthesis of Au nanoparticles stabilized with polyethyleneglycols with different molecular weights from 200 to 8000 Da was carried out. The synthesis was carried out by the method of chemical reduction in an aqueous medium using sodium citrate as a reducing agent. The dependence of the optical properties on the concentration and molar mass of polyethyleneglycol was studied in the obtained samples of Au nanoparticles. The absorption spectra were recorded using an SF-56 optical spectrometer. The studies were carried out in the visible range of the spectrum from 400 to 800 nm. It was found that the type of spectrum, the position of the surface plasmon resonance band and the optical density of the samples of Au nanoparticles stabilized with PEG-8000 with a concentration of 10 and 20% did not undergo significant changes during storage, which characterizes the high aggregate stability of these sols. The dispersed characteristics of these samples of sols of Au nanoparticles were also studied. The studies were carried out using photon-correlation spectroscopy by the method of dynamic light scattering. It is established that an increase in the concentration of the stabilizer leads to an increase in the average hydrodynamic radius of the particles. This fact is associated with an increase in the thickness of the stabilizer layer and with the "stitching" of the polymer layer of Au nanoparticles with the formation of aggregates. Thus, the best result was found in PEG-8000 samples with concentrations of 10 and 20%, since the type of spectrum, the position of the surface plasmon resonance band and the optical density did not undergo significant changes. Based on the data obtained, it can be concluded that the best stabilizer for Au nanoparticles obtained by the citrate method is PEG-8000 with a concentration of at least 10 %. It is important to note that with an increase in the concentration of the stabilizer, the average hydrodynamic radius of the particles increases. This fact is associated with an increase in the thickness of the stabilizer layer and with the "stitching" of Au nanoparticles.

Elevated NTCP expression by an iPSC-derived human hepatocyte maintenance medium enhances HBV infection in NTCP-reconstituted HepG2 cells

Background The sodium taurocholate cotransporting polypeptide (NTCP) is a functional receptor for hepatitis B virus (HBV). NTCP-reconstituted human hepatoma cells support HBV infection, but the infection is suboptimal and no apparent HBV spread has been observed in this system. Results We found that NTCP-reconstituted HepG2 cells were highly susceptible to HBV infection after cells were cultured in a commercial human inducible pluripotent stem cell (iPSC)-derived hepatocyte maintenance medium (HMM). The enhanced HBV infection coincided with increased NTCP expression, and was observed in six different clones of HepG2-NTCP cells. Promoter assays indicated that HMM activated the cytomegalovirus immediate-early (IE) promoter that drives the NTCP expression in the HepG2-NTCP cells. RNA-Seq analysis revealed that HMM upregulated multiple metabolic pathways. Despite highly upregulated NTCP expression by HMM, no obvious HBV spread was observed even in the presence of PEG 8000. Conclusions Our data suggest that this particular medium could be used to enhance HBV infection in NTCP-reconstituted hepatocytes in vitro.

MADURACIÓN Y GERMINACIÓN DE EMBRIONES SOMÁTICOS DE Coffea arabica CV. COLOMBIA

La variedad de café Colombia (Coffea arabica L.) tiene alta resistencia a la roya y buenas características de producción y calidad de taza. Diversos protocolos para la propagación de café mediante embriogénesis somática han sido desarrollados; sin embargo, la tasa de maduración, germinación y conversión es baja. El objetivo de la presente investigación fue determinar el efecto de reguladores del crecimiento y agentes osmóticos en la maduración y germinación de embriones somáticos de café y evaluar, en condiciones ex vitro, las características fisiológicas de las plantas regeneradas. Para su maduración, embriones globulares y torpedo temprano se cultivaron en un medio Yasuda et al. (1985) con distintas concentraciones de ácido abscísico (ABA) (2-4 mg L-1), ácido salicílico (ASA) (1.38 g L-1), ácido giberélico (AG3) (0.7-1.0 mg L-1), ácido indolacético (AIA) (0.3-0.5 mg L-1), polietilenglicol 8000 (PEG 8000) (50-75 g L-1) o sacarosa (50-80 g L-1). Para su germinación, los embriones maduros fueron transferidos a un medio con 0.25 mg L-1 de AIA y 0.25 mg L-1 de 6-bencilaminopurina (BAP). Las plantas se aclimataron y crecieron en perlita:turba:tezontle (1:1:1) o turba:tezontle (2:1) en condiciones de invernadero. El valor máximo de maduración y germinación de los embriones globulares se observó al adicionar 50 g L-1 de sacarosa al medio (81.7 y 53.9 %, respectivamente). Para los embriones torpedo temprano el porcentaje de maduración más alto se registró con 80 g L-1 de sacarosa (19.2 %) o una combinación de 1.0 mg L-1 de AG3 y 0.5 mg L-1 de AIA (45.0 %); sin embargo, los porcentajes de germinación no fueron significativamente distintos del testigo. El potencial osmótico de los embriones no estuvo directamente relacionado con su grado de madurez. La totalidad de las plantas cultivadas en las mezclas de sustratos sobrevivieron y mostraron ser morfológica y fisiológicamente normales bajo condiciones ex vitro.

Silicon Stimulates Plant Growth and Metabolism in Rice Plants under Conventional and Osmotic Stress Conditions

Exogenous silicon (Si) can enhance plant resistance to various abiotic factors causing osmotic stress. The objective of this research was to evaluate the application of 1 and 2 mM Si to plants under normal conditions and under osmotic stress. Morelos A-98 rice seedlings, were treated with 1 and 2 mM SiO2 for 28 d. Subsequently, half of the plants were subjected to osmotic stress with the addition of 10% polyethylene glycol (PEG) 8000; and continued with the addition of Si (0, 1 and 2 mM SiO2) for both conditions. The application of Si under both conditions increased chlorophyll b in leaves, root volume, as well as fresh and dry biomass of roots. Interestingly, the number of tillers, shoot fresh and dry biomass, shoot water content, concentration of total chlorophyll, chlorophyll a/b ratio, and the concentration of total sugars and proline in shoot increased with the addition of Si under osmotic stress conditions. The addition of Si under normal conditions decreased the concentration of sugars in the roots, K and Mn in roots, and increased the concentration of Fe and Zn in shoots. Therefore, Si can be used as a potent inorganic biostimulant in rice Morelos A-98 since it stimulates plant growth and modulates the concentration of vital biomolecules and essential nutrients.

Effect of Seed Coating and PEG-Induced Drought on the Germination Capacity of Five Clover Crops

The effect of coating the seed of clover crops by water absorbing seed process (WASP) technology pelletization on its germination capacity was studied in conditions of diverse drought intensities simulated by different concentrations of polyethylenglycol (PEG) 8000 solution. Drought resistance was monitored in the seed of five fodder clover species: Anthyllis vulneraria L., Medicago lupulina L., Trifolium repens L., Melilotus albus Medik. and Onobrychis viciifolia Scop. In the seed of given plant species, germination capacity was determined along with the share of dead and hard seeds. Although the coating significantly (p < 0.05) affected the drought resistance of seeds, the germination capacity increased only in conditions of milder drought (simulation with PEG: 0.1–0.3 mol). With the increasing intensity of drought induced by higher PEG concentrations (0.4–0.7 mol) the number of germinable seeds demonstrably decreased and the number of dead seeds increased in the coated seed as compared with the uncoated seed. The coated seed can be appropriate for use in M. lupulina, M. albus and T. repens, while the uncoated seed can be used in A. vulneraria and O. viciifolia.

Tuning Cell-free Composition Controls the Time-delay, Dynamics, and Productivity of TX-TL Expression

The composition of TX-TL cell-free expression systems are adjusted by adding macromolecular crowding agents and salts. However, the effects of these cosolutes on the dynamics of individual gene expression processes have not been systematically quantified. Here, we carry out kinetic mRNA and protein level measurements on libraries of genetic constructs using the common cosolutes PEG-8000, Ficoll-400, and magnesium glutamate. By combining these measurements with biophysical modeling, we show that cosolutes have differing effects on transcription initiation, translation initiation, and translation elongation rates with trade-offs between time-delays, expression tunability, and maximum expression productivity. We also confirm that biophysical models can predict translation initiation rates in TX-TL using E. coli lysate. We discuss how cosolute composition can be tuned to maximize performance across different cell-free applications, including biosensing, diagnostics, and biomanufacturing.

Purification and Crystallographic Analysis of a Novel Cold-Active Esterase (HaEst1) from Halocynthiibacter arcticus

This report deals with the purification, characterization, and a preliminary crystallographic study of a novel cold-active esterase (HaEst1) from Halocynthiibacter arcticus. Primary sequence analysis reveals that HaEst1 has a catalytic serine in G-x-S-x-G motif. The recombinant HaEst1 was cloned, expressed, and purified. SDS-PAGE and zymographic analysis were carried out to characterize the properties of HaEst1. A single crystal of HaEst1 was obtained in a solution containing 10% (w/v) PEG 8000/8% ethylene glycol, 0.1 M Hepes-NaOH, pH 7.5. Diffraction data were collected to 2.10 Å resolution with P21 space group. The final Rmerge and Rp.i.m values were 7.6% and 3.5% for 50–2.10 Å resolution. The unit cell parameters were a = 35.69 Å, b = 91.21 Å, c = 79.15 Å, and β = 96.9°.