Water-Molecule-Induced Reversible Fluorescence in a One-Dimensional Mn-Based Hybrid Halide for Anticounterfeiting and Digital Encryption-Decryption.

Hybrid metal halides with reversible transformation of structure and luminescence properties have attracted significant attention in anticounterfeiting. However, their long transition time and slow response rate may hinder the rapid identification of confidential information. Here, a one-dimensional hybrid manganese-based halide, i.e., (C5H11N3)MnCl2Br2·H2O, is prepared and demonstrates the phenomenon of water-molecule-induced reversible photoluminescence transformation. Heating for <40 s induces a dynamic transfer of red-emissive (C5H11N3)MnCl2Br2·H2O to green-emissive (C5H11N3)MnCl2Br2. In addition, the green emission can gradually revert to red emission during a cooling process in a moist environment, demonstrating excellent reversibility. It is found that the water molecule acts as an external stimulus to realize the reversible transition between red and green emission, which also exhibits remarkable stability during repeated cycles. Furthermore, with the assistance of heating and cooling, a complex digital encryption-decryption and an optical "AND" logical gate are achieved, facilitating the development of anticounterfeiting information security.

Negatively charged phospholipids accelerate the membrane fusion activity of the plant-specific insert domain of an aspartic protease.

Various plants use antimicrobial proteins/peptides to resist phytopathogens. In the potato, Solanum tuberosum, the plant-specific insert (PSI) domain of an aspartic protease performs this role by disrupting phytopathogen plasma membranes. However, the mechanism by which PSI selects target membranes has not been elucidated. Here, we studied PSI-induced membrane fusion, focusing on the effects of lipid composition on fusion efficiency. Membrane fusion by the PSI involves an intermediate state whereby adjacent liposomes share their bilayers. We found that increasing the concentration of negatively charged phosphatidylserine (PS) phospholipids substantially accelerated PSI-mediated membrane fusion. NMR data demonstrated that PS did not affect the binding between the PSI and liposomes but had seminal effects on the dynamics of PSI interaction with liposomes. In PS-free liposomes, the PSI underwent significant motion, which was suppressed on PS-contained liposomes. Molecular dynamics simulations showed that the PSI binds to PS-containing membranes with a dominant angle ranging from -31° to 30°, with respect to the bilayer, and is closer to the membrane surfaces. In contrast, PSI is mobile and exhibits multiple topological states on the surface of PS-free membranes. Taken together, our data suggested that PS lipids limit the motion of the anchored PSI, bringing it closer to the membrane surface and efficiently bridging different liposomes to accelerate fusion. As most phytopathogens have a higher content of negatively charged lipids as compared with host cells, these results indicate that the PSI selectively targets negatively charged lipids, which likely represents a way of distinguishing the pathogen from the host.

Coacervates Forming Coexisting Phases on a Mineral Surface.

Minerals played a crucial role in the chemical evolution of small molecules into biopolymers. Yet, it is still not clear how the minerals are related to the formation and the evolution of protocells on early Earth. In this work, using the coacervate formed by quaternized dextran (Q-dextran) and single-stranded oligonucleotides (ss-oligo) as the protocell model, we systematically studied the phase separation of Q-dextran and ss-oligo on the muscovite surface. Serving as rigid and 2D polyelectrolytes, the muscovite surface can be treated by Q-dextran to become negatively charged, neutral, or positively charged. We observed that Q-dextran and ss-oligo form uniform coacervates on naked and neutral muscovite surfaces, while they form biphasic coacervates containing Q-dextran-rich phases and ss-oligo-rich phases on positively or negatively charged muscovite surfaces that were pretreated by Q-dextran. The evolution of the phases is caused by the redistribution of the components as the coacervate touches the surface. Our study indicates that the mineral surface could be a potential driving force for the formation of protocells with hierarchical structures and desirable functions on prebiotic Earth.

Crowded Environment Regulates the Coacervation of Biopolymers via Nonspecific Interactions.

The membrane-less organelles (MLOs) with subcompartments are formed via liquid-liquid phase separation (LLPS) in the crowded cell interior whose background molecules are up to 400 mg/mL. It is still a puzzle how the background molecules regulate the formation, dynamics, and functions of MLOs. Using biphasic coacervate droplets formed by poly(l-lysine) (PLL), quaternized dextran (Q-dextran), and single-stranded oligonucleotides (ss-oligo) as a model of MLO, we online monitored the LLPS process in Bovine Serine Albumin (BSA) solution up to 200.0 mg/mL. Negatively charged BSA is able to form complex or coacervate with positively charged PLL and Q-dextran and thus participates in the LLPS via nonspecific interactions. Results show that BSA effectively regulates the LLPS by controlling the phase distribution, morphologies, and kinetics. With increasing BSA concentration, the spherical biphasic droplets evolve in sequence into phase-inverted flower-like structure, worm-like chains, network structures, and confined coacervates. Each kind of morphology is formed via its own specific growth and fusion pathway. Our work suggests that MLOs could be controlled solely by the crowded environment and provides a further step toward understanding the life process in cell.

Solvent-Free Grinding Synthesis of Hybrid Copper Halides for White Light Emission.

Lead-free metal halides (LMHs) have recently attracted numerous attention in solid-state lighting due to their unique structures and outstanding optoelectronic properties. However, conventional preparation processes with the utilization of toxic organic solvents and high temperatures seem to impede commercial applications of LMHs. In this work, we successfully synthesize Cu+-based metal halides (TMA)3Cu2Br5-xClx (TMA: tetramethylammonium) with high photoluminescence quantum yields (PLQYs) via a solvent-free mechanical grinding method. By changing the ratio of halide ions (Cl- and Br-) in precursors, the emission wavelength of the prepared (TMA)3Cu2Br5-xClx can be tuned from 535 to 587 nm, which are employed as emitters in the fabrication of white-light-emitting diodes (WLEDs). The achieved WLEDs exhibit a high color rendering index value of 84 and standard Commission Internationale de l'Éclairage (CIE) coordinates of (0.324, 0.333). This feasible and solvent-free preparation strategy not only promotes the mass production of LMHs but also highlights the promising potential for efficient solid-state illumination.

Significant performance enhancement of UV-Vis self-powered CsPbBr3 quantum dot-based photodetectors induced by ligand modification and P3HT embedding.

In this work, we report a novel, to the best of our knowledge, strategy to improve the performance of UV-Vis self-powered CsPbBr3 quantum dot (QD) based photodetectors (PDs) by ligand modification and poly(3-hexylthiophene) (P3HT) embedding. Compared with those based on pure QDs, modified PDs show a shortened response time by nearly ten times, and increases of maximum responsivity and specific detectivity by nearly 45 and 97 times, respectively. Such PDs also show a high stability with 90% of the initial photocurrent being maintained even after storage in ambient air without any encapsulation for 30 days.

Coacervate Formed by an ATP-Binding Aptamer and Its Dynamic Behavior under Nonequilibrium Conditions.

Although numerous protocell models have been developed to explore the possible pathway of the origin of life on the early earth, few truly fulfill the roles of the DNA/RNA sequence and ATP molecules, which are keys to cell replication and evolution. The ATP-binding aptamer offers an opportunity to combine sequence and energy molecules. In this work, we choose the coacervate droplet as the protocell model and investigate the interaction of the DNA aptamer, poly(l-lysine)(PLL), and ATP under varying conditions. PLL and aptamers form solid precipitates, which spontaneously transform to coacervate droplets as ATP is introduced. The selective uptake and sequestration of exogenous molecules is achieved by the ATP-containing coacervates. As an electric field is applied to expel ATP, the portion of the droplet deficient in ATP becomes solid. The solid/liquid phase ratio is tunable by varying the electric field strength and excitation time. Together with the vacuolization process, a solid head with a soft mouth periodically opening and closing is created. Moreover, the composite coacervate droplet gradually grows as it is treated with an electric field and cannot recover to the original liquid phase after the power is turned off and replenished with ATP. Our work highlights that the proper integration of the DNA sequence, ATP, and energy input could be a powerful strategy for creating a protocell with certain living features.

CELSR1 variants are associated with partial epilepsy of childhood.

CELSR1 gene, encoding cadherin EGF LAG seven-pass G-type receptor 1, is mainly expressed in neural stem cells during the embryonic period. It plays an important role in neurodevelopment. However, the relationship between CELSR1 and disease of the central nervous system has not been defined. In this study, we performed trios-based whole-exome sequencing in a cohort of 356 unrelated cases with partial epilepsy without acquired causes and identified CELSR1 variants in six unrelated cases. The variants included one de novo heterozygous nonsense variant, one de novo heterozygous missense variant, and four compound heterozygous missense variants that had one variant was located in the extracellular region and the other in the cytoplasm. The patients with biallelic variants presented severe epileptic phenotypes, whereas those with heterozygous variants were associated with a mild epileptic phenotype of benign epilepsy with centrotemporal spikes (BECTS). These variants had no or low allele frequency in the gnomAD database. The frequencies of the CELSR1 variants in this cohort were significantly higher than those in the control populations. The evidence from ClinGen Clinical-Validity Framework suggested a strong association between CELSR1 variants and epilepsy. These findings provide evidence that CELSR1 is potentially a candidate pathogenic gene of partial epilepsy of childhood.

Dynamic Behavior of Complex Coacervates with Internal Lipid Vesicles under Nonequilibrium Conditions.

During the evolution of life on earth, the emergence of lipid membrane-bounded compartments is one of the most enigmatic events. Endosymbiosis has been hypothesized as one of the solutions. In this work, using a coacervate droplet formed by single-stranded oligonucleotides (ss-oligo) and poly(l-lysine) (PLL) as the protocell model, we monitored the uptake of liposomes of different types and studied the dynamic behavior of the resulting composite droplet under the electric field. The coacervate droplet exhibits affinity for the liposomes of varying charges. However, the permeation of liposome is also controlled by electrostatic interactions. Dominated by electrostatic attraction, the positively charged liposome is retained inside the droplet as growing fibrous structures, while the negatively charged liposome is mainly coated on the droplet surface. Permeation and even distribution occur when the liposome and the droplet carry the same charges, or at least one of them is neutral. As an electric field is applied to trigger repetitive cycles of vacuolization in the ss-oligo/PLL droplet, the fibrous structure formed by the positively charged liposome is basically intact, while a new phase is generated together with uneven mass transport as the negatively charged liposome is internalized. Interestingly, the release of daughter droplets with similar components occurs on the droplet containing neutral liposomes. Our work not only provides a step toward creating protocells with hierarchical structures and biofunctions using a biogenetic material via simple mixing but also sheds light on the possible origin of the lipid structure inside a living organism.

Fission and Internal Fusion of Protocell with Membraneless "Organelles" Formed by Liquid-Liquid Phase Separation.

Construction of protocells with hierarchical structures and living functions is still a great challenge. Growing evidence demonstrates that the membraneless organelles, which facilitate many essential cellular processes, are formed by RNA, protein, and other biopolymers via liquid-liquid phase separation (LLPS). The formation of the protocell in the early days of Earth could follow the same principle. In this work, we develop a novel coacervate-based protocell containing membraneless subcompartments via spontaneous liquid-liquid phase separation by simply mixing single-stranded oligonucleotides (ss-oligo), quaternized dextran (Q-dextran), and poly(l-lysine) (PLL) together. The resulting biphasic droplet, with PLL/ss-oligo phase being the internal subcompartments and Q-dextran/ss-oligo phase as the surrounding medium, is capable of sequestering and partitioning biomolecules into distinct regions. When the droplet is exposed to salt or Dextranase, the surrounding Q-dextran/ss-oligo phase will be gradually dissociated, resulting in the chaotic movement and fusion of internal subcompartments. Besides, the external electric field at a lower strength can drive the biphasic droplet to undergo a deviated circulation concomitant with the fusion of localized subcompartments, while a high-strength electric field can polarize the whole droplet, resulting in the release of daughter droplets in a controllable manner. Our study highlights that liquid-liquid phase separation of biopolymers is a powerful strategy to construct hierarchically structured protocells resembling the morphology and functions of living cells and provides a step toward a better understanding of the transition mechanism from nonliving to living matter under prebiotic conditions.

PEGylated gene carriers in serum under shear flow.

The behaviour of drug/gene carriers in the blood stream under shear is still a puzzle. In this work, using the complexes formed by 21 bp DNA and poly(ethylene glycol)-b-poly(l-lysine) (PEG-PLL) of varying PEG lengths, we studied the dynamic behaviour of the complexes in the presence of fetal bovine serum (FBS) and under flow at different shear rates, a condition mimicking the internal physical environment of blood vessels. The PEG5k-PLL/DNA complex possesses a dense DNA/PLL core and a loose PEG5k protecting layer. The PEGylated DNA complexes exhibit multiple responses to external shear in the presence of FBS. The loose PEG5k layer is firstly disturbed at a shear rate below 30 s-1. The exposure of the charged core to the environment results in a secondary aggregation of the complex with FBS. The size of the aggregate is limited to a certain range as the shear rate increases to 50 s-1. The dense DNA/PLL core starts to withstand the shear force as the shear rate reaches 500 s-1. The reorganization of the core to accommodate more serum molecules leads to tertiary aggregation of the complexes. If PEG cannot form a valid layer around the complex, as in PEG2k-PLL/DNA, the complex forms an aggregate even without shear, and the first shear dependent region is missing. If the PEG layer is too stable around the complex, as in PEG10k-PLL/DNA, no tertiary aggregation occurs. The mechanism of shear on the behaviour of delivery particles in serum helps to design gene carriers with high efficacy.

Mass Transport in Coacervate-Based Protocell Coated with Fatty Acid under Nonequilibrium Conditions.

Construction of protocell models from prebiotically plausible components to mimic the basic features or functions of living cells is still a challenge. In this work, we prepare a hybrid protocell model by coating sodium oleate on the coacervate droplet constituted by poly(l-lysine) and oligonucleotide and investigate the transport of different molecules under electric field. Results show that sodium oleate forms a layered viscoelastic membrane on the droplet surface, which is selectively permeable to small, polar molecules, such as oligolysine. As the droplet is stimulated at 10 V cm-1, the oleate membrane slips along the direction of electric field while maintaining its integrity. Most of the molecules are still excluded under such conditions. As repetitive cycles of vacuolization occur at 20 V cm-1, all molecules are internalized and sequestrated in the droplet through their specific pathways except enzyme, which anchors in the oleate membrane and is immune to electric field. Cascade enzymatic reactions are then carried out, and the product generated from the membrane exhibits a time-dependent concentration gradient across the droplet. Our work makes a step toward the nonequilibrium functionalization of synthetic protocells capable of biomimetic operations.

Inward Budding and Endocytosis of Membranes Regulated by de Novo Designed Peptides.

Protein-mediated endocytosis of membrane is a key event in biological system. The mechanism, however, is still not clear. Using a de novo designed bola-type peptide KKKLLLLLLLLKKK (K3L8K3) as a protein mimic, we studied how it induced giant unilamellar vesicle (GUV) to form inward buds or endocytosis at varying conditions. Results show that the inward budding is initiated as the charged lipids are neutralized by K3L8K3, which results in a negative spontaneous curvature. If the charged lipids have unsaturated tails, the buddings are slim fibrils, which can further wrap into a spherical structure. In the case of saturated charged lipids, the buddings are rigid tubules, stable in the studied time period. The unsaturated lipid to saturated lipid ratio in the mother membrane is another key parameter governing the shape and dynamics of the buds. A complete endocytosis is observed when K3L8K3 is attached with a hydrophobic moiety, suggesting that hydrophobic interaction helps the buds to detach from the mother membrane. The molecules in the surrounding medium, such as negatively charged oligonucleotides, are engulfed into the GUV via endocytosis pathway induced by K3L8K3. Our study provides a novel strategy for illustrating the endocytosis mechanism by using peptides of simple sequence.

Electric field-induced circulation and vacuolization regulate enzyme reactions in coacervate-based protocells.

Artificial protocells operating under non-equilibrium conditions offer a new approach to achieve dynamic features with life-like properties. Using coacervate micro-droplets comprising polylysine (PLL) and a short single-stranded oligonucleotide (ss-oligo) as a membrane-free protocell model, we demonstrate that circulation and vacuolization can occur simultaneously inside the droplet in the presence of an electric field. The circulation is driven by electrohydrodynamics and applies specifically to the major components of the protocell (PLL and ss-oligo). Significantly, under low electric fields (E = 10 V cm-1) the circulation regulates the movement of the vacuoles, while high levels of vacuolization produced at higher electric fields can deform or reshape the circulation. By taking advantage of the interplay between vacuolization and circulation, we achieve dynamic localization of an enzyme cascade reaction at specific droplet locations. In addition, the spatial distribution of the enzyme reaction is globalized throughout the droplet by tuning the coupling of the circulation and vacuolization processes. Overall, our work provides a new strategy to create non-equilibrium dynamic behaviors in molecularly crowded membrane-free synthetic protocells.

Shear Effects on Stability of DNA Complexes in the Presence of Serum.

The behavior of nanocarriers, even though they are well-defined at equilibrium conditions, is unpredictable in living system. Using the complexes formed by plasmid DNA (pDNA) and K20 (K: lysine), protamine, or polylysine (PLL) as models, we studied the dynamic behavior of gene carriers in the presence of fetal bovine serum (FBS) and under different shear rates, a condition mimicking the internal physical environment of blood vessels. Without shear, all the positively charged complexes bind to the negatively charged proteins in FBS, leading to the formation of large aggregates and even precipitates. The behaviors are quite different under shear. The shear generates two effects: a mechanical force to break down the complex into smaller size particles above a critical shear rate and a stirring effect leading to secondary aggregation of complexes below the critical shear rate. In the studied shear rate from 100 to 3000 s-1, the mechanical force plays a key role in K20/pDNA and protamine/pDNA, while the stirring effect is dominant in PLL/pDNA. A model study shows that the interfacial tension, the chain density, and the elasticity of the complexes determine their responsiveness to shear force. This study is helpful to understand the fate of drug/gene carriers under physiological conditions. It also gains insight in designing drug/gene carriers with desirable properties for in vivo applications.

Enzymatic activity inside a DNA/peptide complex.

The mutual interaction between enzymes and their environments plays a key role in various life processes. In this study, using the complexes formed by salmon DNA and a de novo designed peptide, Ac-RRRRRRRRRGALGLPGKGGGLQRLTALDGR-NH2 (abbreviated as RR-30), as a model, we studied the activity of collagenase encapsulated inside the complex. Collagenase is able to cleave RR-30 at a LG/LP site, generating two shorter length peptides, which decreases the stability of the complex. Results show that the complex dissociates with time in the presence of collagenase. The dissociation rate is linearly proportional to the collagenase concentration. On the other hand, the collagenase activity is severely deteriorated inside the complex, where only 1/3 of the enzyme is active. We attribute it to the electrostatic interaction and hydrophobic interaction between collagenase and the components of the complex. Therefore, the mutual interaction determines the structure and kinetics of the DNA/peptide complex.

Effects of Structural Flexibility on the Kinetics of DNA Y-Junction Assembly and Gelation.

The kinetics of DNA assembly is determined not only by temperature but also by the flexibility of the DNA tiles. In this work, the flexibility effect was studied with a model system of Y-junctions, which contain single-stranded thymine (T) loops in the center. It was demonstrated that the incorporation of a loop with only one thymine prominently improved the assembly rate and tuned the final structure of the assembly, whereas the incorporation of a loop of two thymines exhibited the opposite effect. These observations could be explained by the conformation adjustment rate and the intermotif binding strength. Increasing DNA concentration hindered the conformational adjustment rate of DNA strands, leading to the formation of hydrogels in which the network was connected by ribbons. Therefore, the gel can be treated as a metastable state during the phase transition.

Effects of chain flexibility on the properties of DNA hydrogels.

The effect of chain rigidity on the mechanic properties of DNA hydrogels was studied. Counterintuitively, the hydrogel formed by mainly flexible chains exhibited better stability, stretchability, and much mechanical properties than the hydrogel containing only rigid chains. Calculations showed that the crosslinking ratio in the hydrogel formed by flexible chains was about twice that of the hydrogel formed by rigid chains under the same conditions. We attributed this to the ease of conformational adjustment of flexible chains. Incorporation of 25% rigid chains further improved the performance of DNA hydrogel by shrinking the pore size and tuning its distribution.

Relationship between peptide structure and antimicrobial activity as studied by de novo designed peptides.

As fundamental components in innate immunity, antimicrobial peptides (AMPs) hold great potentials in the treatment of persistent infections involving slow-growing or dormant bacteria in which, selective inhibition of prokaryotic bacteria in the context of eukaryotic cells is not only an essential requirement, but also a critical challenge in the development of antimicrobial peptides. To identify the sequence and structural properties critical for antimicrobial activity, a series of peptides varying in sequence, length, hydrophobicity/charge ratio, and secondary structure, were designed and synthesized. Their antimicrobial activities were then tested using Escherichia coli and HEK293 cells, together with several index activities against model membrane, including liposome leakage, fusion, and aggregation. While no evident correlation between the antimicrobial activity and the property of the peptides was observed, common activities against model membrane were nevertheless identified for the active antimicrobial peptides: mediating efficient membrane leakage, negligible membrane fusion and liposome aggregation. Therefore, in addition to identifying one highly active antimicrobial peptide, our study further sheds light on the design principle for these molecules.

Axial growth and fusion of liposome regulated by macromolecular crowding and confinement.

The endomembrane system, including the endoplasmic reticulum, Golgi apparatus, lysosomes, and endosomes, is located in the crowded intracellular environment. An understanding of the cellular structure and functions requires knowledge of how macromolecular crowding and confinement affect the activity of membrane and its proteins. Using negatively charged liposome and the peptide K3L8K3 as a model system, we studied the aggregation behavior of liposome in a matrix of polyacrylamide and hyaluronic acid. Without matrix, the liposomes form spherical aggregates in the presence of K3L8K3. However, they orient in one dimension and fuse into a tube up to 40 µm long in the matrix. The growth of the tube is via end-to-end connection. This anisotropic growth is mainly due to the macromolecular confinement provided by the polymer network. The study of the interactions between liposome and peptide in the crowded environment helps to reveal the mechanism of membrane-related processes in vivo.