Charged Protein Research Articles

The Donnan equilibrium is still valid in high-volume HDF.

Clinical studies have shown that hemodiafiltration reduces morbidity and mortality of dialysis patients compared to hemodialysis alone. This is attributed to its superior middle molecule clearance compared to standard hemodialysis. However, doubts arose as to whether a high convective flux through the dialyzer membrane has an influence on the equilibrium concentration of small ions, especially that of sodium. Due to the presence of negatively charged impermeable proteins on the blood side, the Gibbs-Donnan effect leads to an asymmetric distribution of membrane permeable ions on both sides of the membrane. In thermodynamic equilibrium, the concentrations of those ions can easily be calculated. However, the convective fluid flow leads to deviations from thermodynamic equilibrium. In this article, the effect of a convective flow on the ion distribution across a semipermeable membrane is analyzed in a theoretical model. Starting from the extended Nernst-Planck equation, including diffusive, convective, and electrostatic effects, a set of differential equations is derived. An approximate solution for flow speeds up to 0.1 ms-1 as well as a numerical solution are given. The results show that in any practical dialysis setting the convective flow has negligible influence on the electrolyte concentrations.

Driving Forces in the Formation of Biocondensates of Highly Charged Proteins: A Thermodynamic Analysis of the Binary Complex Formation

A thermodynamic analysis of the binary complex formation of the highly positively charged linker histone H1 and the highly negatively charged chaperone prothymosin α (ProTα) is detailed. ProTα and H1 have large opposite net charges (−44 and +53, respectively) and form complexes at physiological salt concentrations with high affinities. The data obtained for the binary complex formation are analyzed by a thermodynamic model that is based on counterion condensation modulated by hydration effects. The analysis demonstrates that the release of the counterions mainly bound to ProTα is the main driving force, and effects related to water release play no role within the limits of error. A strongly negative Δcp (=−0.87 kJ/(K mol)) is found, which is due to the loss of conformational degrees of freedom.

Effects of mono- and divalent- ions and their strength on foaming properties of infant formula protein model system

Metal ions may affect the foaming properties of infant formula products that including milk protein during reconstitution. In this study, an infant formula protein model system (IFPMS) composed of both whey protein and casein was constructed, and the effect of mono (Na+, K+) and divalent (Mg2+, Ca2+) ions and their strength (0–100 mM) on foaming properties was investigated. The mono ions had no significantly impact on foaming capacity (FC), MgCl2 slightly increase FC and reaching a maximum of 116.09 ± 7.46% at 25 mM, whereas CaCl2 significantly decreased FC from 105.78 ± 7.42% to 83.14 ± 6.75% as 0-25 mM (p < 0.05). Foam stability (FS) was significantly improved as all salts above 10 mM (p < 0.05). The dramatically decreased net charge and larger protein aggregates (> 25 mM) of IFPMS with divalent ions indicating their stronger charge-shielding effect, which caused slow adsorption of protein and increased surface tension. Multiple spectral results confirmed that salts enhanced protein interactions via inter-molecular and form a viscoelastic layer to stabilize foam. Absorbed protein and SDS-PAGE analysis revealed that MgCl2 promoted protein adsorption, while CaCl2 reduced casein in the foam phase. Correlation analysis further implied that the FC showed highly significant positive with absorbed protein, solubility, and β-turn (p ≤ 0.01), and FS was significantly correlated with secondary structure and viscosity (p ≤ 0.05). This study may provide useful information for an in-depth understanding of the foaming properties of infant formula.

Nanoetched Stainless Steel Architecture Enhances Cell Uptake of Biomacromolecules and Alters Protein Corona Abundancy.

Nanotexture on biocompatible surfaces promotes cell adhesion and proliferation. High aspect ratio nanoachitecture serves as an ideal interface between implant materials and host cells that is well-suited for localized therapeutic delivery. Despite this potential, nanotextured surfaces have not been widely applied for biomacromolecule delivery. Here, we employed a low-cost, industrially relevant nanoetching process to modify the surface of biocompatible stainless steel 316 (SS316L), creating nanotextured SS316L (NT-SS316L) as a material for intracellular biomacromolecule delivery. As biomacromolecule cargoes are adsorbed to the steel and ultimately would be used in protein-rich environments, we performed serum protein corona analysis on unmodified SS316L and NT-SS316L using tandem mass spectrometry. We observed an increase in proteins associated with cell adhesion on the surface of NT-SS316L compared to that of SS316L, supporting literature reports of enhanced adhesion on nanotextured materials. For delivery to adherent cells, a "hard corona" of model biomacromolecule cargoes including superfolder green fluorescent protein (sfGFP) charge variants, cytochrome c, and siRNA was adsorbed on NT-SS316L to assess delivery. Nanotextured surfaces enhanced cellular biomacromolecule uptake and delivered cytosolic-functional proteins and nucleic acids through energy-dependent endocytosis. Collectively, these findings indicate that NT-SS316L holds potential as a surface modification for implants to achieve localized drug delivery for a variety of biomedical applications.

Adenosine Triphosphate Inhibits Cold-Responsive Aggregation.

Adenosine triphosphate (ATP), ubiquitous in all living organisms, is conventionally recognized as a fundamental energy currency essential for a myriad of cellular processes. While its traditional role in energy metabolism requires only micromolar concentrations, the cellular content of ATP has been found to be significantly higher at the millimolar level. Recent studies have attempted to correlate this higher concentration of ATP with its nonenergetic role in maintaining protein homeostasis, leaving the investigation of ATP's nontrivial activities in biology an open question. Here, by coupling computer simulations and experiments, we uncover new insights into ATP's role as a cryoprotectant against cold-salt stress, highlighting the necessity for higher cellular ATP concentrations. We present direct evidence at charged silica interfaces, demonstrating ATP's ability to restore native intersurface interactions disrupted by combined cold-salt stress, thereby inhibiting cold-responsive aggregation in high-salt conditions. ATP desorbs salt cations from negatively charged surfaces through predominant interactions between ATP and the salt cations. Although the mode of ATP's action remains unchanged with temperature, the extent of interaction scales with temperature, requiring less ATP activity at lower temperatures, justifying the reason for reduction in cellular ATP content due to the cold effect, reported in previous experimental studies. The trend observed in inorganic nanostructures is recurrent and robustly transferable to charged protein interfaces. A thorough comparison of ATP's cryoprotective activity with traditionally known biological cryoprotectants (glycine and betaine) reveals ATP's greater efficiency. In retrospect, our findings highlight ATP's additional biological role in cryopreservation, expanding its potential biomedical applications by offering effective protection of cells from cryoinjuries and avoiding the significant challenges associated with the toxicity of organic cryoprotectants.

Characterization of a Charged Biomimetic Lipid Membrane for Unique Antifouling Effects against Clinically Relevant Matrices in Biosensing.

Clinically relevant matrices such as human blood and serum can cause substantial interference in biosensing measurements, severely compromising the effectiveness of the sensors. We report the characterization of a positively charged lipid membrane that has demonstrated unique features to suppress the nonspecific signal for antifouling effects by using SPR, fluorescence recovery after photobleaching (FRAP), and MALDI-TOF-MS. The ethylphosphocholine (EPC) lipid membrane proved to be exceptionally effective at reducing irreversible interactions from human serum on a Protein A surface. The membrane formation conditions and their effects on membrane fluidity and mobility were characterized for understanding the antifouling functions when various capture molecules were immobilized. Specifically, EPC lipid membranes on a Protein A substrate appear to exhibit a strong interaction, likely through the electrostatic effect with the negatively charged proteins that resulted in a stable hydration layer. The strong interaction also limited lipid mobility, contributing to a robust, protective interface that remained undamaged in undiluted serum. Tailoring a surface with antifouling lipid membranes allows for a range of biosensing applications in highly complex biological media.

Inhibition of RNA splicing triggers CHMP7 nuclear entry, impacting TDP-43 function and leading to the onset of ALS cellular phenotypes.

Amyotrophic lateral sclerosis (ALS) is linked to the reduction of certain nucleoporins in neurons. Increased nuclear localization of charged multivesicular body protein 7 (CHMP7), a protein involved in nuclear pore surveillance, has been identified as a key factor damaging nuclear pores and disrupting transport. Using CRISPR-based microRaft, followed by gRNA identification (CRaft-ID), we discovered 55 RNA-binding proteins (RBPs) that influence CHMP7 localization, including SmD1, a survival of motor neuron (SMN) complex component. Immunoprecipitation-mass spectrometry (IP-MS) and enhanced crosslinking and immunoprecipitation (CLIP) analyses revealed CHMP7's interactions with SmD1, small nuclear RNAs, and splicing factor mRNAs in motor neurons (MNs). ALS induced pluripotent stem cell (iPSC)-MNs show reduced SmD1 expression, and inhibiting SmD1/SMN complex increased CHMP7 nuclear localization. Crucially, overexpressing SmD1 in ALS iPSC-MNs restored CHMP7's cytoplasmic localization and corrected STMN2 splicing. Our findings suggest that early ALS pathogenesis is driven by SMN complex dysregulation.

A biodegradable lipid nanoparticle delivers a Cas9 ribonucleoprotein for efficient and safe in situ genome editing in melanoma

The development of melanoma is closely related to Braf gene, which is a suitable target for CRISPR/Cas9 based gene therapy. CRISPR/Cas9-sgRNA ribonucleoprotein complexes (RNPs) stand out as the safest format compared to plasmid and mRNA delivery. Similarly, lipid nanoparticles (LNPs) emerge as a safer alternative to viral vectors for delivering the CRISPR/Cas9-sgRNA gene editing system. Herein, we have designed multifunctional cationic LNPs specifically tailored for the efficient delivery of Cas9 RNPs targeting the mouse Braf gene through transdermal delivery, aiming to treat mouse melanoma. LNPs are given a positive charge by the addition of a newly synthesized polymer, deoxycholic acid modified polyethyleneimine (PEI-DOCA). Positive charge enables LNPs to be delivered in vivo by binding to negatively charged cell membranes and proteins, thereby facilitating efficient skin penetration and enhancing the delivery of RNPs into melanoma cells for gene editing purposes. Our research demonstrates that these LNPs enhance drug penetration through the skin, successfully delivering the Cas9 RNPs system and specifically targeting the Braf gene. Cas9 RNPs loaded LNPs exert a notable impact on gene editing in melanoma cells, significantly suppressing their proliferation. Furthermore, in mice experiments, the LNPs exhibited skin penetration and tumor targeting capabilities. This innovative LNPs delivery system offers a promising gene therapy approach for melanoma treatment and provides fresh insights into the development of safe and effective delivery systems for Cas9 RNPs in vivo. Statement of significanceCRISPR/Cas9 technology brings new hope for cancer treatment. Cas9 ribonucleoprotein offers direct genome editing, yet delivery challenges persist. For melanoma, transdermal delivery minimizes toxicity but faces skin barrier issues. We designed multifunctional lipid nanoparticles (LNPs) for Cas9 RNP delivery targeting the Braf gene. With metal microneedle pretreatment, our LNPs effectively edited melanoma cells, reducing Braf expression and inhibiting tumor growth. Our study demonstrates LNPs' potential for melanoma therapy and paves the way for efficient in vivo Cas9 RNP delivery systems in cancer therapy.

Phase behavior and interaction of strong polyelectrolyte dextran sulfate and whey protein isolation: Effects of pH, protein/polysaccharide ratio, and salt addition

The strong polyelectrolyte dextran sulfate (DS) is an anionic polysaccharide with a high negative charge, characterized by high stability and pH independence. DS and whey protein isolate (WPI) were selected to study the specific effects of highly negatively charged polysaccharides on the phase behavior and interaction of WPI/DS complexes (1 % w/v) under varying external conditions (pH, WPI:DS ratio, and salt addition). The phase diagrams, zeta potential, and laser confocal scanning microscopy measurements indicated that the WPI/DS complexes did not dissociate even at pH 1 due to the pH independence of DS. The exclusion volume effect of DS promoted WPI self-aggregation at high salt concentrations, which inhibited acidification-induced dissociation. Isothermal titration calorimetry indicated that the WPI/DS interaction is a spontaneous exothermic reaction driven by both enthalpy and entropy changes due to electrostatic interactions. This study provides valuable information on the interactions between highly negatively charged polysaccharides and proteins.

Advancing Protein Analysis: A Low-Pressure Drift Tube Orbitrap Mass Spectrometer for Ultraviolet Photodissociation-Based Structural Characterization.

Owing to its ability to generate extensive fragmentation of proteins, ultraviolet photodissociation (UVPD) mass spectrometry (MS) has emerged as a versatile ion activation technique for the structural characterization of native proteins and protein complexes. Interpreting these fragmentation patterns provides insight into the secondary and tertiary structures of protein ions. However, the inherent complexity and diversity of proteins often pose challenges in resolving their numerous conformations. To address this limitation, we combined UVPD-MS with drift tube ion mobility, offering potential to acquire conformationally selective MS/MS information. A low-pressure drift tube (LPDT) Orbitrap mass spectrometer equipped with 193 nm UVPD capabilities enables the analysis of protein conformers through the analysis of arrival time distributions (ATDs) of individual fragment ions. ATDs of fragment ions are compared for different backbone cleavage sites of the protein or different precursor charge states to give information about regions of potential folding or elongation. This integrated platform offers promise for advancing our understanding of protein structures in the gas phase.

Monitoring Binding of Protamine with DNA Using Protein Charge Transfer Spectra.

In this work, novel intrinsic electronic absorption (250-400 nm) with a molar extinction coefficient of 752 M-1cm-1 at 250 nm, arising from photoinduced electron transfer involving charged amino acid side chains and the polypeptide backbone, along with luminescence (300-500 nm) with quantum yield of 0.011 from subsequent charge recombination, was observed in salmon sperm Protamine (PRM). The absorption of PRM was attributed to the previously identified Peptide Backbone-to-Side chain Charge Transfer (PBS-CT) from the polypeptide backbone to the abundant cationic headgroups of Arginine in PRM, while the luminescence was believed to originate from charge recombination within the charge-separated excited states of PRM. Remarkably, since Arg is the only charged residue in the PRM sequence, the PRM Protein Charge Transfer Spectra (ProCharTS) is both totally and uniquely Arg specific. Interestingly, the peak of PRM luminescence emission spectrum was independent of the excitation wavelength, unlike other proteins such as human serum albumin, displaying unconventional luminescence. Aggregation-induced effects on PRM absorbance and luminescence were ruled out, as both PRM absorbance and luminescence increase maintained linearity with increasing concentration in the 25-150 μM range. Nucleoprotamine complex formation, resulting from the binding of PRM with calf-thymus genomic DNA (gDNA), was monitored through increased scattering by the nucleoprotamine complex, a decrease in gDNA/PRM absorbance, a decrease in gDNA/PRM ellipticity, and shifts of nucleoprotamine complex band upon agarose gel electrophoresis. Upon binding with gDNA (700 μM base pair concentration), PRM ProCharTS absorbance at 260 nm decreased by 72%. This decrease was attributed to the formation and subsequent precipitation of nucleoprotamine complex upon PRM-gDNA binding. The application of ProCharTS absorbance to indirectly monitor DNA-protein binding in a label-free approach was thus demonstrated.

Statistical Thermodynamic Description of Self-Assembly of Large Inclusions in Biological Membranes.

Recent studies revealed anomalous underscreening in concentrated electrolytes, and we suggest that the underscreened electrostatic forces between membrane proteins play a significant role in the process of self-assembly. In this work, we assumed that the underscreened electrostatic forces compete with the thermodynamic Casimir forces induced by concentration fluctuations in the lipid bilayer, and developed a simplified model for a binary mixture of oppositely charged membrane proteins with different preference to liquid-ordered and liquid-disordered domains in the membrane. In the model, like macromolecules interact with short-range Casimir attraction and long-range electrostatic repulsion, and the cross-interaction is of the opposite sign. We determine energetically favored patterns in a system in equilibrium with a bulk reservoir of the macromolecules. Different patterns consisting of clusters and stripes of the two components and of vacancies are energetically favorable for different values of the chemical potentials. Effects of thermal flutuations at low temperature are studied using Monte Carlo simulations in grand canonical and canonical ensembles. For fixed numbers of the macromolecules, a single two-component cluster with a regular pattern coexists with dispersed small one-component clusters, and the number of small clusters depends on the ratio of the numbers of the molecules of the two components. Our results show that the pattern formation is controlled by the shape of the interactions, the density of the proteins, and the proportion of the components.

In situ formation of biomolecular condensates as intracellular drug reservoirs for augmenting chemotherapy.

Biomolecular condensates, which arise from liquid-liquid phase separation within cells, may provide a means of enriching and prolonging the retention of small-molecule drugs within cells. Here we report a method for the controlled in situ formation of biomolecular condensates as reservoirs for the enrichment and retention of chemotherapeutics in cancer cells, and show that the approach can be leveraged to enhance antitumour efficacies in mice with drug-resistant tumours. The method involves histones as positively charged proteins and doxorubicin-intercalated DNA strands bioorthogonally linked via a click-to-release reaction between trans-cyclooctene and tetrazine groups. The reaction temporarily impaired the phase separation of histones in vitro, favoured the initiation of liquid-liquid phase separation within cells and led to the formation of biomolecular condensates that were sufficiently large to be retained within tumour cells. The controlled formation of biomolecular condensates as drug reservoirs within cells may offer new options for boosting the efficacies of cancer therapies.

Remarkable Insensitivity of Protein Diffusion to Protein Charge.

Friction to translational diffusion of ionic particles in polar liquids should scale linearly with the squared ion charge, according to standard theories. Substantial slowing of translational diffusion is expected for proteins in water. In contrast, our simulations of charge mutants of green fluorescent proteins in water show remarkable insensitivity of the translational diffusion constant to protein's charge in the range of charges between -29 and +35. The friction coefficient is given as a product of the force variance and the memory function relaxation time. We find remarkably accurate equality between the variance of the electrostatic force and the negative cross-correlation of the electrostatic and van der Waals forces. The charge invariance of the diffusion constant is a combined effect of the force variance and relaxation time invariances with the protein charge. The temperature dependence of the protein diffusion constant is highly non-Arrhenius, with a fragile-to-strong crossover at the glass transition.

An innovative method for fractionating the milk fat globule membrane and the major buttermilk proteins

Buttermilk (BM) is a by-product of butter manufacturing that shares a similar composition to skim milk but contains a higher concentration of phospholipids due to milk fat globule membrane (MFGM) fragments released during cream churning. Despite its high added-value components, MFGM is underutilized mainly due to the challenge in separating its components from the other constituents of buttermilk. Our work has focused on the use of calcium phosphate particles, called hydroxyapatite (HA), to extract and separate the different components from buttermilk. This study investigated the impact of various physicochemical parameters (temperature, ionic strength, and pH) on the adsorption of a mixture of micellar casein (CM), whey proteins, and MFGM from BM onto HA surfaces. A box-Behnken design was utilized to predict optimal physicochemical conditions for selectively separating components in buttermilk. Ionic strength (IS) played a crucial role, with low IS having a minimal positive influence and high IS negatively influencing adsorption. pH influenced adsorption by altering protein and HA surface charges. The optimal conditions for adsorption were an IS of 100 mM for a pH of 7, for 90 % of CM, 37 % of β-lactoglobulin, 11 % of α-lactalbumin and 7 % of MFGM. Desorption experiments using citrate and alkaline pH showed promising results for CM recovery, with citrate efficiently releasing the adsorbed CM. Additionally, MFGM were separated from whey proteins through selective precipitation at a pH of 4.0, allowing sedimentation of MFGM (100 %) while keeping whey proteins soluble. The presence of β-lactoglobulin in the protein fraction of MFGM (31 %) and CM (37 %) was attributed to pasteurization, which denatured a portion of β-lactoglobulin and led to its attachment to MFGM surfaces and interaction with CM. This protocol produces a nearly pure fraction of MFGM with a simple method which is suitable for industrial production, representing significant valorization for the dairy sector.

Light-Activated Molecular Purification (LAMP) of Recombinant Proteins.

The production of recombinant proteins has become a focal point in biotechnology, with potential applications in catalysis, therapeutics, and diagnostics. Before their application, these proteins undergo cumbersome downstream processing, including multiple resin-based chromatography steps (ion exchange or affinity-based) to isolate the protein of interest from host cell proteins, which are more abundant. These methods often involve (1) nonspecific binding of host cell proteins onto the resin, (2) a trial and error approach in determining elution conditions for the protein of interest, and (3) complex functionalization of the resin. These processes are also further supplemented with additional processing steps including buffer exchange through dialysis or desalting. Despite the prevalence and need for proteins, challenges persist in optimizing elution conditions and minimizing downstream processing steps, which contribute to the overall cost, impeding their translation into the market. To address these challenges, there has been a growing interest in stimuli-responsive purification systems, which allow for precise control and modulation of the purification process for protein recovery by altering their properties or behavior in response to specific external conditions, such as heat, light, or chemicals. We have developed a light-activated molecular purification (LAMP) system, a stimuli-responsive chromatography technique where the purification of recombinant proteins is triggered by light. We employed a photocleavable protein (PhoCl1) that binds specifically to Ni-NTA resin through a hexa-histidine tag at its N-terminus. We harnessed the ability of PhoCl1 to undergo photocleavage into two fragments for the development of LAMP. To demonstrate LAMP, the protein of interest (POI) is genetically fused to the C-terminus of PhoCl1. The exposure to 405 nm light (1.5 mW cm-2 for 12 h) results in the release of POI into the supernatant. We showcased the potential of LAMP by purifying highly charged green fluorescent proteins and an enzyme, riboflavin kinase. Our custom-built violet light LED setup achieved more than 50% light-induced photocleavage of the fusion constructs, resulting in the release of more than 30% of the POI into the supernatant, with the remainder retained within the resin. All the proteins purified using LAMP were more than 90% pure. Moreover, the comparison of the riboflavin kinase purified through LAMP and the traditional chromatography (Ni-NTA affinity method) revealed no significant changes in the activity levels. These highlight the broad potential of LAMP in providing a facile, yet robust stimuli-responsive protein purification technique, which leverages the potential of light to purify the proteins and overcome the limitations of current conventional chromatography systems.

Micronuclear collapse from oxidative damage.

Chromosome-containing micronuclei are a hallmark of aggressive cancers. Micronuclei frequently undergo irreversible collapse, exposing their enclosed chromatin to the cytosol. Micronuclear rupture catalyzes chromosomal rearrangements, epigenetic abnormalities, and inflammation, yet mechanisms safeguarding micronuclear integrity are poorly understood. In this study, we found that mitochondria-derived reactive oxygen species (ROS) disrupt micronuclei by promoting a noncanonical function of charged multivesicular body protein 7 (CHMP7), a scaffolding protein for the membrane repair complex known as endosomal sorting complex required for transport III (ESCRT-III). ROS retained CHMP7 in micronuclei while disrupting its interaction with other ESCRT-III components. ROS-induced cysteine oxidation stimulated CHMP7 oligomerization and binding to the nuclear membrane protein LEMD2, disrupting micronuclear envelopes. Furthermore, this ROS-CHMP7 pathological axis engendered chromosome shattering known to result from micronuclear rupture. It also mediated micronuclear disintegrity under hypoxic conditions, linking tumor hypoxia with downstream processes driving cancer progression.

Conformational and Structural Characterization of Knotted Proteins.

Knotted proteins are fascinating natural biomolecules whose backbones entangle themselves in a knot. Their particular knotted configurations provide them with a wide range of topological features. However, their folding/unfolding mechanisms, stability, and function are poorly understood. In the present work, native trapped ion mobility spectrometry-mass spectrometry (TIMS-MS) was used for characterizing structural features of two model knotted proteins: a Gordian 52 knot ubiquitin C-terminal hydrolase (UCH) and a Stevedore 61 knot (α-haloacid dehalogenase, DehI). Experimental results showed structural transitions of UCH and DehI as a function of solution composition (0-50% MeOH) and temperature (T ∼20-95 °C). An increase in the protein charge states and collision cross sections (∼2750-8750 Å2 and ∼3250-15,385 Å2 for UCH and DehI, respectively) with the solution organic content (OC) and temperature suggested a three-step unfolding pathway with at least four structural transitions. Results also showed that the integrity of the UCH knot core was more resistant to thermal unfolding when compared to DehI; however, both knot cores can be disrupted with the increase in the solution OC. Additional enzymatic digestion experiments using carboxypeptidase Y combined with molecular dynamics simulations showed that the knot core was preserved between Glu20 and Glu188 and Arg89 and His304 residues for UCH and DehI, respectively, where disruption of the knot core led to structural collapse followed by unfolding events. This work highlights the potential of solution OC and temperature studies combined with native TIMS-MS for the comprehensive characterization of knotted proteins to gain a better understanding of their structural transitions.

A “like-zwitterionic” forward osmosis membranes with electrostatic deposition of aluminosilicate nanotubes for resisting positively and negatively charged protein and oil contaminant

Membrane fouling has always been a huge obstacle to the development of membrane separation technology, including forward osmosis (FO) technology. In this work, polyamide (PA) thin-film composite (TFC) membranes were simply immersed in aluminosilicate nanotubes (ANTs) dispersions to obtain thin-film nanocomposite (TFN) FO membranes. With the help of the opposite surface charge of ANTs and PA layer, and the representative “peak-valley” structure of PA TFC membrane, ANTs are stably bound to the surface of TFC membrane by electrostatic adsorption and physical confinement. The TFN membrane has greatly enhanced hydration ability due to the rich hydrophilic hydroxyl groups of ANTs, “like-zwitterionic” surface composition and rough nanostructure morphology, resulting in good underwater superoleophobicity. Combined with the water transport channel formed by its nanoporous structure, the surface anti-protein adhesion and anti-oil–water emulsion adhesion can be achieved without affecting the water flux. It is worth noting that the surface potential of TFN membrane is close to neutral due to charge neutralization. This avoids the electrostatic interaction between the membrane surface and the contaminant, and further achieves a universal anti-fouling ability that is not affected by protein or emulsion charge, and the irreversible adsorption is close to zero. In addition, the rigid ANTs coating exhibits excellent stability and reusability under long-term use, and the water flux after cleaning can be restored to nearly 99.9%. In general, this method of using ANTs deposition to improve the antifouling performance of PA based TFC membrane is simple, efficient, stable, and easy scalable. This method is not only limited to FO membrane, but also can be extended to PA based reverse osmosis and nanofiltration membranes, which has great practical application potential.

Influence of amphiphilic chitosan-deoxycholic acid-polyacrylic acid copolymer on activity, dynamics, and structure of human lysozyme and pyrazinamidase

Cationic chitosan, a naturally degradable and biocompatible polysaccharide, has shown considerable promise as a drug delivery system. However, its limited solubility in water at neutral and alkaline pH has restricted its broader application. To overcome this challenge, various chemical modification strategies have been explored to enhance the solubility of chitosan. One such strategy involves the development of chitosan-deoxycholic acid-polyacrylic acid (CDP) copolymers, which are novel amphiphilic derivatives of chitosan that exhibit excellent water solubility and self-assembly properties. Herein, we investigated the potential of CDP as an anionic carrier for two model proteins: the positively charged Lysozyme and the negatively charged Pyrazinamidase. The objective of this investigation is to elucidate the role of electrostatic interactions between the copolymer and the proteins during the drug encapsulation process. A combined approach utilizing experimental techniques and molecular dynamics (MD) simulations was used to investigate the interactions between the polymer and the proteins, as well as any potential structural changes to the proteins. The results suggest contrasting binding mechanisms for the two proteins: Lysozyme being entrapped within the cationic polymer network and Pyrazinamidase being adsorbed onto the surface of the polymeric assemblies. Binding energy calculations revealed favorable physical interactions between CDP and Lysozyme, while Pyrazinamidase exhibited unfavorable binding. These findings demonstrated that CDP may serve as a promising carrier for positively charged drugs due to its favorable interactions and potential entrapment within the polymer network. The favorable interactions observed between CDP and Lysozyme suggest that CDP could be further explored as a delivery system for positively charged therapeutic proteins.