Self-Emergent Protocells Generated in an Aqueous Solution with Binary Macromolecules through Liquid-Liquid Phase Separation

Recently, liquid–liquid phase separation (LLPS) has attracted considerable attention among researchers in the life sciences as a plausible mechanism for the generation of microstructures inside cells. LLPS occurs through multiple nonspecific interactions and does not always require a lock-and-key interaction with a binary macromolecular solution. The remarkable features of LLPS include the non-uniform localization and concentration of solutes, resulting in the ability to isolate certain chemical systems and thereby parallelize multiple chemical reactions within the limited space of a living cell. We report that, by using the macromolecules, poly(ethylene glycol) (PEG) and dextran, that exhibit LLPS in an aqueous solution, cell-sized liposomes are spontaneously formed therein in the presence of phospholipids. In this system, LLPS is generated through the depletion effect of macromolecules. The results showed that cell-like microdroplets entrapping DNA wrapped by a phospholipid layer emerge in a self-organized manner.     

Specific Spatial Localization of Actin and DNA in a Water/Water Microdroplet: Self‐Emergence of a Cell‐Like Structure

The effect of binary hydrophilic polymers on a pair of representative bio‐macromolecules in a living cell has been examined. The results showed that these bio‐macromolecules exhibited specific localization in cell‐sized droplets that were spontaneously formed through water/water microphase segregation under crowding conditions with coexisting polymers. In these experiments, a simple binary polymer system with poly(ethylene glycol) (PEG) and dextran (DEX) was used. Under the conditions of microphase segregation, DNA was entrapped within cell‐sized droplets rich in DEX. Similarly, F‐actin, linearly polymerized actin, was entrapped specifically within microdroplets rich in DEX, whereas G‐actin, a monomeric actin, was distributed evenly inside and outside these droplets. This study has been extended to a system with both F‐actin and DNA, and it was found that DNA molecules were localized separately from aligned F‐actin proteins to create microdomains inside microdroplets, reflecting the self‐emergence of a cellular morphology similar to a stage of cell division.     

FTIR Spectroscopy to Reveal Lipid and Protein Changes Induced on Sperm by Capacitation: Bases for an Improvement of Sample Selection in ART

Although being a crucial step for Assisted Reproduction Technologies (ART) success, to date sperm selection is based only on morphology, motility and concentration characteristics. Considering the many possible alterations, there is a great need for analytical approaches allowing more effective sperm selections. The use of Fourier Transform Infrared (FTIR) may represent an interesting possibility, being able to reveal many macromolecular changes in a single measurement in a nondestructive way. As a proof of concept, in this observational study, we used a FTIR approach to reveal features related to sperm quality and chemical changes promoted by in vitro capacitation. We found indication that α-helix content is increased in capacitated sperm, while high percentages of the β-structures seem to correlate to poor-quality spermatozoa. The most interesting observation was related to the lipid composition, when measured as CH₂/CH₃ vibrations (ratio 2853/2870), which resulted in being strongly influenced by capacitation and well correlated with sperm motility. Interestingly, this ratio is higher than 1 in infertile samples, suggesting that motility is related to sperm membranes stiffness and lipid composition. Although further analyses are requested, our results support the concept that FTIR can be proposed as a new smart diagnostic tool for semen quality assessment in ART.     

Solution‐NMR Characterization of Outer‐Membrane Protein A from E. coli in Lipid Bilayer Nanodiscs and Detergent Micelles

X‐ray crystallography and solution NMR of detergent‐reconstituted OmpA (outer membrane protein A from E. coli) had shown that this protein forms an eight‐stranded transmembrane β‐barrel, but only limited information was obtained for the extracellular loops. In NMR studies of OmpA in two different detergent micelles, “NMR‐invisible” amino acid residues in‐between the extracellular loops and the β‐barrel prevented complete structural characterization. Here, we show that this NMR‐invisible ring around the β‐barrel of OmpA is also present in lipid bilayer nanodiscs and in mixed micelles with a third detergent, thus suggesting that the implicated rate processes have a functional role rather than representing an artifact of the protein reconstitution. In addition to sequence‐specific NMR assignments for OmpA in the nanodiscs, the present results are based on a protocol of micro‐coil TROSY‐ and CRINEPT‐type NMR diffusion measurements for studying the hydrodynamic properties and the foldedness of [²H,¹⁵N]‐labeled membrane proteins in nanodiscs. This protocol can be applied under conditions closely similar to those used for NMR structure determinations or crystallization trials.     

应用遗传算法求解多相化学反应体系的相稳定性及平衡组成

基于吉布斯自由能最小法原理,研究用遗传算法求解复杂化学体系的平衡问题。将相数和相态未知体系的平衡计算问题分解为两步最优化问题:假定相态数目下的吉布斯自由能最小化问题和相稳定性判据问题。通过序贯加入新相和用遗传算法交替求解两个最优化子问题,可得到最终体系的相数、相态和平衡组成。对合成甲醇和酯化反应两个多相复杂体系进行了的计算,结果验证了本研究方法的有效性。     

Effects of (R)- and (S)-α-Hydroxylation of Acyl Chains in Sphingosine,Dihydrosphingosine, and Phytosphingosine Ceramides on Phase Behavior and Permeability of Skin Lipid Models

Ceramides (Cers) with α-hydroxylated acyl chains comprise about a third of all extractable skin Cers and are required for permeability barrier homeostasis. We have probed here the effects of Cer hydroxylation on their behavior in lipid models comprising the major SC lipids, Cer/free fatty acids (C ₁₆-C ₂₄)/cholesterol, and a minor component, cholesteryl sulfate. Namely, Cers with (R)-α-hydroxy lignoceroyl chains attached to sphingosine (Cer AS), dihydrosphingosine (Cer AdS), and phytosphingosine (Cer AP) were compared to their unnatural (S)-diastereomers and to Cers with non-hydroxylated lignoceroyl chains attached to sphingosine (Cer NS), dihydrosphingosine (Cer NdS), and phytosphingosine (Cer NP). By comparing several biophysical parameters (lamellar organization by X-ray diffraction, chain order, lateral packing, phase transitions, and lipid mixing by infrared spectroscopy using deuterated lipids) and the permeabilities of these models (water loss and two permeability markers), we conclude that there is no general or common consequence of Cer α-hydroxylation. Instead, we found a rich mix of effects, highly dependent on the sphingoid base chain, configuration at the α-carbon, and permeability marker used. We found that the model membranes with unnatural Cer (S)-AS have fewer orthorhombically packed lipid chains than those based on the (R)-diastereomer. In addition, physiological (R)-configuration decreases the permeability of membranes, with Cer (R)-AdS to theophylline, and increases the lipid chain order in model systems with natural Cer (R)-AP. Thus, each Cer subclass makes a distinct contribution to the structural organization and function of the skin lipid barrier.     

生育酚/CO2、油酸甲酯/CO2二元高压相平衡测定与关联

利用气液相交替循环装置结合重量法,分别测定了生育酚/CO2、油酸甲酯/CO2两个二元体系在温度为313 15～343 15K,压力为10～20MPa的高压相平衡数据,根据气液相中CO2的p-V-T关系以及相平衡特征,计算出生育酚、油酸甲酯在超临界CO2中的溶解度及其在气液两相中的分配系数。同时将实验测定值与相关文献值进行了比较。最后,运用BP人工神经网络对实验数据进行了关联,计算值与实验值吻合较好。