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Responsive polymeric nanoparticles are an exciting field of study, yet little is known about the temperature sensitivity of those based on tertiary amine methacrylate block copolymers.

Structures and functions of eukaryotic cells with an outer permeable membrane, motility, a cytoskeleton, biomolecules diffusion, and functional organelles can be imitated by a multi-compartmentalized and large-sized protocell containing various synthetic organelles. Herein, two kinds of artificial organelles with stimuli-trigged regulation ability, glucose oxidase-(GOx)-loaded pH-responsive polymersomes A (GOx-Psomes A) and urease-loaded pH-responsive polymersomes B (Urease-Psomes B) for probing biomimetic pH homeostasis, and a pH-sensor (Dextran-FITC) are encapsulated into proteinosomes by Pickering emulsion method. Thus, the polymersomes-in-proteinosome system is realized. Alternating input-regulation of fuels (glucose or urea) outside the protocell penetrates the membrane of proteinosomes and enters into GOx-Psomes A and Urease-Psomes B to produce chemical\biological signals (gluconic acid or ammonia) resulting in pH-feedback loops (pH jump and pH drop). This will counteract the catalytic “switch on” or “switch off” of enzyme-loaded Psomes A and B owing to their different pH-responsive membranes. Thus, the Dextran-FITC promotes a controlled cytosolic spatial organization and the detection of slight pH fluctuations in the lumen of protocells. Overall, this approach shows heterogeneous polymersome-in-proteinosome architectures with sophisticated features such as induced input-regulated pH changes mediated by negative/positive feedback in loops and cytosolic pH self-monitoring, requirements strictly needed in an advanced protocell design.

Most sophisticated biological functions and features of cells are based on self-organization, and the coordination and connection between their cell organelles determines their key functions. Therefore, spatially ordered and controllable self-assembly of polymersomes to construct clusters to simulate complex intracellular biological functions has attracted widespread attention. Here, we present a simple one-step copper-free click strategy to cross-link nanoscale pH-responsive and photo-cross-linked polymersomes (less than 100 nm) to micron-level clusters (more than 90% in 0.5-2 μm range). Various influencing factors in the clustering process and subsequent purification methods were studied to obtain optimal clustered polymeric vesicles. Even when polymeric vesicles separately loaded with different enzymes (glucose oxidase and myoglobin) are coclustered, the overall permeability of the clusters can still be regulated through tuning the pH values on demand. Compared with simple blending of those enzyme-loaded polymersomes, the rate of enzymatic cascade reaction increased significantly due to the interconnected complex microstructure established. The connection of catalytic nanocompartments into clusters confining different enzymes of a cascade reaction provides an excellent platform for the development of artificial systems mimicking natural organelles or cells.

Liver fibrosis affects millions of people worldwide and is rising vastly over the past decades. With no viable therapies available, liver transplantation is the only curative treatment for advanced diseased patients. Excessive accumulation of aberrant extracellular matrix (ECM) proteins, mostly collagens, produced by activated hepatic stellate cells (HSCs), is a hallmark of liver fibrosis. Several studies have suggested an inverse correlation between collagen-I degrading matrix metalloproteinase-1 (MMP-1) serum levels and liver fibrosis progression highlighting reduced MMP-1 levels are associated with poor disease prognosis in patients with liver fibrosis. We hypothesized that delivery of MMP-1 might potentiate collagen degradation and attenuate fibrosis development. In this study, we report a novel approach for the delivery of MMP-1 using MMP-1 decorated polymersomes (MMPsomes), as a surface-active vesicle-based ECM therapeutic, for the treatment of liver fibrosis. The storage-stable and enzymatically active MMPsomes were fabricated by a post-loading of Psomes with MMP-1. MMPsomes were extensively characterized for the physicochemical properties, MMP-1 surface localization, stability, enzymatic activity, and biological effects. Dose-dependent effects of MMP-1, and effects of MMPsomes versus MMP-1, empty polymersomes (Psomes) and MMP-1 + Psomes on gene and protein expression of collagen-I, MMP-1/TIMP-1 ratio, migration and cell viability were examined in TGFβ-activated human HSCs. Finally, the therapeutic effects of MMPsomes, compared to MMP-1, were evaluated in vivo in carbon-tetrachloride (CCl4)-induced early liver fibrosis mouse model. MMPsomes exhibited favorable physicochemical properties, MMP-1 surface localization and improved therapeutic efficacy in TGFβ-activated human HSCs in vitro. In CCl4-induced early liver fibrosis mouse model, MMPsomes inhibited intra-hepatic collagen-I (ECM marker, indicating early liver fibrosis) and F4/80 (marker for macrophages, indicating liver inflammation) expression. In conclusion, our results demonstrate an innovative approach of MMP-1 delivery, using surface-decorated MMPsomes, for alleviating liver fibrosis.

Even the most advanced protein-polymer conjugate therapeutics do not eliminate antibody-protein and receptor-protein recognition. Next-generation bioconjugate drugs will need to replace stochastic selection with rational design to select desirable levels of protein-protein interaction while retaining function. The "Holy Grail" for rational design would be to generate functional enzymes that are fully catalytic with small molecule substrates while eliminating interaction between the protein surface and larger molecules. Using chymotrypsin, an important enzyme that is used to treat pancreatic insufficiency, we have designed a series of molecular chimeras with varied grafting densities and shapes. Guided by molecular dynamic simulations and next-generation molecular chimera characterization with asymmetric flow field-flow fractionation chromatography, we grew linear, branched, and comb-shaped architectures from the surface of the protein by atom-transfer radical polymerization. Comb-shaped polymers, grafted from the surface of chymotrypsin, completely prevented enzyme inhibition with protein inhibitors without sacrificing the ability of the enzyme to catalyze the hydrolysis of a peptide substrate. Asymmetric flow field-flow fractionation coupled with multiangle laser light scattering including dynamic light scattering showed that nanoarmor designed with comb-shaped polymers was particularly compact and spherical. The polymer structure significantly increased protein stability and reduced protein-protein interactions. Atomistic molecular dynamic simulations predicted that a dense nanoarmor with long-armed comb-shaped polymer would act as an almost perfect molecular sieve to filter large ligands from substrates. Surprisingly, a conjugate that was composed of 99% polymer was needed before the elimination of protein-protein interactions.

In recent years, chitosan has attracted considerable interest in many fields due to its sufficient charge density under biological, non-hazardous conditions. Since chitosan originates from natural resources and has two different monomer units, its characterization must be carried out in a goal-oriented and precise manner. This work focuses on the characterization of chitosans most important parameters – solubility, crystallinity, degree of deacetylation (DD) and molecular weight – in a simple and convenient way. The DD was determined using Nuclear Magnetic Resonance spectroscopy (NMR), Particle Charge Detection (PCD), Fourier Transform Infrared spectroscopy (FTIR), CHN elemental analysis (CHN-EA) and conductometric/potentiometric titration with special attention to its physical state as solid or liquid. Investigation of DD by FTIR was successfully determined by calculating peak heights, peak areas and peak deconvolution from a linear combination of Gaussian and Lorentzian functions. Asymmetrical flow field flow fractionation with light scattering detection (AF4-LS) was applied in order to calculate molar masses and radii. In addition, pH-potentiometric titrations demonstrated a reproducible displacement of the point of zero charge (PZC) in form of a hysteresis depending on the titration direction. The DD affects the crystallinity, which was determined by deconvolution of the crystalline and amorphous domains.

Feedback-controlled detection of subtle changes of extracellular biomolecules as known from cells is also needed in protocells. Artificial organelles, located in protocells, detect the small variation in pH which is triggered by different amounts of invading glucose, converted by glucose-oxidase into gluconic acid. The approach paves the way for using pH fluctuations-detecting artificial organelles in the lumen of protocells.

In this extended special feature to celebrate the 35th anniversary edition of LCGC Europe, leading figures from the separation science community explore contemporary trends in separation science and identify possible future developments.

The intramolecular chain collapse of linear precursors with systematic variation of molar mass and ligation group density (5, 15, and 30 mol %) into single-chain nanoparticles (SCNPs) was studied by two different separation approaches. The efficiency of size exclusion chromatography with quadruple detection (SEC-D4) was compared to asymmetrical field flow fractionation hyphenated to quintuple detection (AF4-D5) in organic solvent. The application of the unique combination of advanced detection to different separation principles opens up the opportunity to critically evaluate the determination of molar masses and different types of radii for an in-depth understanding of the structural properties affected by the internal folding process. This is achieved by a detailed comparison of assets, drawbacks, and limitations of these approaches based on the systematical screening of different chain lengths and sizes of the precursors and the SCNPs. Furthermore, an alternative strategy for quantitative determination of intramolecular ligation density by a combination of AF4 and UV detection is introduced.

Dehydropolymerisation of methylamine borane (H3B⋅NMeH2) using the well‐known iron amido complex [(PNP)Fe(H)(CO)] (PNP=N(CH2CH2PiPr2)2) (1) gives poly(aminoborane)s by a chain‐growth mechanism. In toluene, rapid dehydrogenation of H3B⋅NMeH2 following first‐order behaviour as a limiting case of a more general underlying Michaelis–Menten kinetics is observed, forming aminoborane H2B=NMeH, which selectively couples to give high‐molecular‐weight poly(aminoborane)s (H2BNMeH)n and only traces of borazine (HBNMe)3 by depolymerisation after full conversion. Based on a series of comparative experiments using structurally related Fe catalysts and dimethylamine borane (H3B⋅NMe2H) polymer formation is proposed to occur by nucleophilic chain growth as reported earlier computationally and experimentally. A silyl functionalised primary borane H3B⋅N(CH2SiMe3)H2 was studied in homo‐ and co‐dehydropolymerisation reactions to give the first examples for Si containing poly(aminoborane)s., Getting poly(aminoborane)s: The well‐known Fe amido complex [(PNP)Fe(H)(CO)] (PNP=N(CH2CH2PiPr2)2) is a potent catalyst for the selective formation of poly(aminoborane)s from methylamine borane and a SiMe3‐substituted analogue. Mechanistic studies support a chain‐growth mechanism with polymer formation by nucleophilic attack at the terminus of a growing B−N chain.

Synthetic platforms for mimicking artificial organelles or for designing multivalent protein therapeutics for targeting cell surface, extracellular matrix, and tissues are in the focus of this study. Furthermore, the availability of a multi-functionalized and stimuli-responsive carrier system is required that can be used for sequential in situ and/or post loading of different proteins combined with post-functionalization steps. Until now, polymersomes exhibit excellent key characteristics to fulfill those requirements, which allow specific transport of proteins and the integration of proteins in different locations of polymeric vesicles. Herein, different approaches to fabricate multivalent protein-loaded, pH-responsive, and pH-stable polymersomes are shown, where a combination of therapeutic action and targeting can be achieved, by first choosing two model proteins such as human serum albumin and avidin. Validation of the molecular parameters of the multivalent biohybrids is performed by dynamic light scattering, cryo-TEM, fluorescence spectroscopy, and asymmetrical flow-field flow fractionation combined with light scattering techniques. To demonstrate targeting functions of protein-loaded polymersomes, avidin post-functionalized polymersomes are used for the molecular recognition of biotinylated cell surface receptors. These versatile protein-loaded polymersomes present new opportunities for designing sophisticated biomolecular nanoobjects in the field of (extracellular matrix) protein therapeutics.

While personalized therapy bears an enormous potential in cancer therapy, the development of flexible, tailorable delivery systems remains challenging. Here, we present a "tool-kit" of various avidin-based bioconjugates (BCs) for the preparation of personalized delivery systems. Corresponding BCs were synthesized using the self-assembly of avidin with various biotinylated ligands, such as one cationic glycodendrimer for dendriplex adsorption and two functional ligands for imaging (glycodendrimers with DOTA or NOTA units) or targeting (biotinylated PEG decorated with ligands). Substituting antibodies for targeting small molecules were coupled to biotin-PEG compounds for addressing the folate receptor (FR), epidermal growth factor receptor (EGFR), and prostate-specific membrane antigen (PSMA). After successful characterization and proof of good storage and redispersion properties of BCs, cytotoxicity assays and first in vivo imaging studies with 99mTc-complexing bioconjugates provide evidence that these BCs and their avidin analogues can be used as tool-kit components in theranostic systems for personalized medicine.

The cross-linking of caseins with microbial transglutaminase (mTGase) was investigated using asymmetric flow field flow fractionation in combination with static and dynamic light scattering detections (AF4-MALS-DLS). This approach allows determining molar mass, molecular size, scaling properties, and apparent densities of casein aggregates prior to and after enzymatic treatment. The results show that, as a consequence of non-covalent interactions, casein molecules associate to form elongated aggregates with a molar mass ( M w ) ranging from approx. 4 × 105 to 1.2 × 106 g mol−1, and a mean radius of gyration ( R g , z ) of approx. 16 nm. Upon enzymatic treatment with mTGase, casein aggregates become more compact as M w increases but R g decreases with ongoing cross-linking reaction. In contrast, the hydrodynamic radius ( R h ) distribution determined by online DLS is rather unaffected by enzymatic cross-linking, indicating that mTGase cross-links casein molecules mainly within distinct aggregates. Furthermore, extensive enzymatic treatments with mTGase change the molecular shapes of casein aggregates towards more compact and denser spherical structures.

Exosomes are emerging as ideal drug delivery vehicles due to their biological origin and ability to transfer cargo between cells. However, rapid clearance of exogenous exosomes from the circulation as well as aggregation of exosomes and shedding of surface proteins during storage limit their clinical translation. Here, we demonstrate highly controlled and reversible functionalization of exosome surfaces with well-defined polymers that modulate the exosome’s physiochemical and pharmacokinetic properties. Using cholesterol-modified DNA tethers and complementary DNA block copolymers, exosome surfaces were engineered with different biocompatible polymers. Additionally, polymers were directly grafted from the exosome surface using biocompatible photo-mediated atom transfer radical polymerization (ATRP). These exosome polymer hybrids (EPHs) exhibited enhanced stability under various storage conditions and in the presence of proteolytic enzymes. Tuning of the polymer length and surface loading allowed precise control over exosome surface interactions, cellular uptake, and preserved bioactivity. EPHs show fourfold higher blood circulation time without altering tissue distribution profiles. Our results highlight the potential of precise nanoengineering of exosomes toward developing advanced drug and therapeutic delivery systems using modern ATRP methods.

International audience; To mimic organelles and cells and to construct next-generation therapeutics, asymmetric functionalization and location of proteins for artificial vesicles is thoroughly needed to emphasize the complex interplay of biological units and systems through spatially separated and spatiotemporal controlled actions, release, and communications. For the challenge of vesicle (= polymersome) construction, the membrane permeability and the location of the cargo are important key characteristics that determine their potential applications. Herein, an in situ and post loading process of avidin in pH-responsive and photo-cross-linked polymersomes is developed and characterized. First, loading efficiency, main location (inside, lumen, outside), and release of avidin under different conditions have been validated, including the pH-stable presence of avidin in polymersomes’ membrane outside and inside. This advantageous approach allows us to selectively functionalize the outer and inner membranes as well as the lumen with several bio(macro)molecules, generally suited for the construction of asymmetrically functionalized artificial organelles. In addition, a fluorescence resonance energy transfer (FRET) effect was used to study the permeability or uptake of the polymersome membrane against a broad range of biotinylated (macro)molecules (different typology, sizes, and shapes) under different conditions.

The eukaryotic cell is a smart compartment containing an outer permeable membrane, a cytoskeleton, and functional organelles, presenting part structures for life. The integration of membrane-containing artificial organelles (=polymersomes) into a large microcompartment is a key step towards the establishment of exquisite cellular biomimetics with different membrane properties. Herein, an efficient way to construct a hierarchical multicompartment composed of a hydrogel-filled proteinosome hybrid structure with an outer homogeneous membrane, a smart cytoskeleton-like scaffold, and polymersomes is designed. Specially, this hybrid structure creates a micro-environment for pH-responsive polymersomes to execute a desired substance transport upon response to biological stimuli. Within the dynamic pH-stable skeleton of the protein hydrogels, polymersomes with loaded PEGylated insulin biomacromolecules demonstrate a pH-responsive reversible swelling-deswelling and a desirable, on-demand cargo release which is induced by the enzymatic oxidation of glucose to gluconic acid. This stimulus responsive behavior is realized by tunable on/off states through protonation of the polymersomes membrane under the enzymatic reaction of glucose oxidase, integrated in the skeleton of protein hydrogels. The integration of polymersomes-based hybrid structure into the proteinosome compartment and the stimuli-response on enzyme reactions fulfills the requirements of eukaryotic cell biomimetics in complex architectures and allows mimicking cellular transportation processes.

Artificial vesicles made of block copolymers, so-called polymersomes, represent a versatile chassis for the creation of functionalized nanocompartments with a wide range of biotechnological applications. The specific application depends on the biomolecules – usually proteins – that are positioned in the interior, in the membrane or on the surface of the vesicles. However, not all added proteins are integrated into the vesicles during the usual manufacturing processes of polymersomes. Excess proteins must therefore be removed. The separation techniques currently used for this, however, are associated with decisive disadvantages, such as damaged vesicles, long process times, or small sample volumes that can be processed. To overcome these drawbacks, we investigated the applicability of Capto™ Core 700 resin for polymersome purification. Polymersomes were not damaged or otherwise affected by passage through the column verified by hollow fiber flow field flow fractionation technique. Using three proteins with divergent physico-chemical properties as examples, it was demonstrated that different types of unentrapped proteins were efficiently removed from polymersome dispersions. The dynamic binding capacities in the presence of polymersomes varied between 9.5 and 16.5 mg per mL resin for the proteins applied. The technique can be used for small and large sample volumes alike. In addition, it can be used without special laboratory equipment. This adds a new and easy-to-use purification method for polymer vesicles to the repertoire that will also facilitate the large-scale production of functionalized polymersomes.

Selective isolation of DNA is crucial for applications in biology, bionanotechnology, clinical diagnostics and forensics. We herein report a smart methanol-responsive polymer (MeRPy) that can be programmed to bind and separate single- as well as double-stranded DNA targets. Captured targets are quickly isolated and released back into solution by denaturation (sequence-agnostic) or toehold-mediated strand displacement (sequence-selective). The latter mode allows 99.8% efficient removal of unwanted sequences and 79% recovery of highly pure target sequences. We applied MeRPy for the depletion of insulin, glucagon, and transthyretin cDNA from clinical next-generation sequencing (NGS) libraries. This step improved the data quality for low-abundance transcripts in expression profiles of pancreatic tissues. Its low cost, scalability, high stability and ease of use make MeRPy suitable for diverse applications in research and clinical laboratories, including enhancement of NGS libraries, extraction of DNA from biological samples, preparative-scale DNA isolations, and sorting of DNA-labeled non-nucleic acid targets., Krieg et al. describe a methanol responsive polymer that can capture complementary DNA using grafted oligonucleotides. They successfully demonstrate its efficacy with simultaneous and sequence-specific isolation of three target genes (cDNA) from clinical NGS libraries with high efficiency. This method is fast, effective, scalable, modular, and versatile.

Temporal activation of biological processes by visible light and subsequent return to an inactive state in the absence of light is an essential characteristic of photoreceptor cells. Inspired by these phenomena, light-responsive materials are very attractive due to the high spatiotemporal control of light irradiation, with light being able to precisely orchestrate processes repeatedly over many cycles. Herein, it is reported that light-driven proton transfer triggered by a merocyanine-based photoacid can be used to modulate the permeability of pH-responsive polymersomes through cyclic, temporally controlled protonation and deprotonation of the polymersome membrane. The membranes can undergo repeated light-driven swelling-contraction cycles without losing functional effectiveness. When applied to enzyme loaded-nanoreactors, this membrane responsiveness is used for the reversible control of enzymatic reactions. This combination of the merocyanine-based photoacid and pH-switchable nanoreactors results in rapidly responding and versatile supramolecular systems successfully used to switch enzymatic reactions ON and OFF on demand.

In this paper, we report on the use of amorphous lignin, a waste by-product of the paper industry, for the production of high performance carbon fibers (CF) as precursor with improved thermal stability and thermo-mechanical properties. The precursor was prepared by blending of lignin with polyacrylonitrile (PAN), which was previously dissolved in an ionic liquid. The fibers thus produced offered very high thermal stability as compared with the fiber consisting of pure PAN. The molecular compatibility, miscibility, and thermal stability of the system were studied by means of shear rheological measurements. The achieved mechanical properties were found to be related to the temperature-dependent relaxation time (consistence parameter) of the spinning dope and the diffusion kinetics of the ionic liquids from the fibers into the coagulation bath. Furthermore, thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), and dynamic mechanical tests (DMA) were utilized to understand in-depth the thermal and the stabilization kinetics of the developed fibers and the impact of lignin on the stabilization process of the fibers. Low molecular weight lignin increased the thermally induced physical shrinkage, suggesting disturbing effects on the semi-crystalline domains of the PAN matrix, and suppressed the chemically induced shrinkage of the fibers. The knowledge gained throughout the present paper allows summarizing a novel avenue to develop lignin-based CF designed with adjusted thermal stability. publishedVersion

Internationale forskningsresultater viser, at lærere på tværs af uddannelsestrin møder udfordringer i deres arbejde med at få elever til at bevæge sig fra en faktapræget reproducerende tilnærmning til kundskabsudvikling til en mere kritisk reflekterende tilnærmning. Hensigten med mit projekt er at identifisere og forstå nogle af disse udfordringer i tilknytning til undervisning i samfundsfaglige temaer i norsk videregående skole. Målet er samtidig at synliggøre, hvordan læreres forståelse af kritisk refleksion, kundskab og kundskabsudvikling er en vigtig præmisleverandør for, hvad kritisk refleksion kan være i skolen. Indgangsvinkelen til projektet er følgende problemstilling: Hvad karakteriserer læreres erfaringer med arbejdet med udvikling af elevers kritiske reflekterende evner i samfundsfaglige temaer i norsk videregående skole? Projektets analytiske perspektiver omhandler forskellige forståelser af og tilnærmninger til kritisk refleksion som kundskab og kompetence. I diskusionkapitlet inddrager jeg dessuden perspektiver på professionsudøvelse og lærerens professionelle pædagogiske handlingsrum. Projektet er gennemført som en kvalitativ studie med interview af fem lærere i norsk videregående skole. Datamaterialet er analyseret gennem en induktiv metode med fokus på kategorisering af semantiske relationer. Resultaterne viser, at de elementer som indrammer deltagernes forståelse af sit eget pædagogiske arbejdsrum i forbindelse med undervisning med fokus på kritisk refleksion kan knyttes til henholdsvis skolen som institution, skolen om organisation og elevernes forudsætninger som overordnede momenter. Resultaterne viser også, hvordan den enkelte deltagers forståelse af og positionering i forhold til elementerne fungerer som betingelse for, hvordan kritisk refleksion kan fremdyrkes i skolen, samt hvordan dette kan ses i sammenhæng med, at der eksisterer forskellige forståelser af kritisk refleksion. Diskusionskapitlet synliggør særligt, hvordan deltagernes positionering i forhold til henholdsvis elevernes manglende faglige ballast og mødet mellem skolens og elevens virkelighed bliver en udfordrende rammebetingelse for, hvad kritisk refleksion kan være i skolen. Results from international research show that teachers across educational levels face challenges when trying to move students from a fact-based, reproductive approach to an approach focusing on knowledge development and critical reflectiveness. The purpose of my project is to identify and understand some of these challenges related to teaching social science subjects in Norwegian high school. The aim is also to visualize how teachers' understanding of critical reflection, knowledge and knowledge development is an important premise for what critical reflection can be in school. The approach to the project is the following thesis question: What characterizes teachers’ experiences when working on developing students' critical reflective skills in social science topics in Norwegian high school? The project's analytical perspectives deal with different understandings and approaches to critical reflection as knowledge and as competence. In the discussion chapter, perspectives on professional practice and the teacher's professional pedagogical space are included. The project was conducted as a qualitative study with interviews of five teachers in Norwegian high school. The data material is analyzed through an inductive method focusing on the categorization of semantic relations. The results show that the elements that frame the participants’ understanding of their own pedagogical workspace in connection with teaching with a focus on critical reflection can be linked to the school as an institution, the school as an organization and the students' qualifications as main elements. The results also show how the individual participant's understanding of and positioning in relation to the elements function as a condition for how critical reflection can be cultivated in school, and furthermore, how this can be seen in correlation with different understandings of critical reflection. The discussion chapter highlights in particular how the participants’ positioning in relation to, respectively, the students' lack of academic ballast and the meeting between the school and the student's reality, becomes a challenging framework condition for what critical reflection can be in school. Masteroppgave i pedagogikk IKT PED396 MAPS-PEDIK

The dinuclear zirconocene chloride complex 1 is a highly active precatalyst for the dehydropolymerisation of methylamine borane. Comparison with mononuclear Zr chlorides and related dinuclear complexes suggests that the nature of the bridging motif is essential for the unique reactivity of 1.

A novel approach for the integration of π-conjugated polymers (CPs) into DNA-based nanostructures is presented. Using the controlled Kumada catalyst-transfer polycondensation, well-defined thiophene-based polymers with controllable molecular weight, specific end groups, and water-soluble oligoethylene glycol-based side chains were synthesized. The end groups were used for the easy but highly efficient click chemistry-based attachment of end-functionalized oligodeoxynucleotides (ODNs) with predesigned sequences. As demonstrated by surface plasmon resonance spectroscopy, the prepared block copolymers (BCPs), P3(EO)3T-b-ODN, comprising different ODN lengths and specific or repetitive sequences, undergo specific hybridization with complementary, thiol-functionalized ODNs immobilized on a gold surface. Furthermore, the site-specific attachment of the BCPs to DNA origami structures is studied. We demonstrate that a nanoscale object, that is, a single BCP with a single ODN handle, can be directed and bound to th...

This study describes new mechanistic insights in the sequential polyassociation of streptavidin with biotinylated poly(ethyleneimine) glycopolymers and biotinylated PEGylated folic acid components for the preparation of biohybrid structures (BHS) for controlled targeting experiments. Characterization of the BHS revealed that during the formation and postfunctionalization of BHS, reversible dissociation and reassociation processes occur. The BHS are stable over weeks after finalizing the equilibrium-driven polyassociation process. Cellular uptake studies showed that this sequential polyassociation involving biotinylated PEGylated folic acid components does not lead to enhanced cellular uptake of the resulting BHS. In contrast, polyplexes, containing small interfering RNA and bioconjugates (1:1 molar ratio between biotinylated glycopolymer and monomeric streptavidin-lectin fusion protein), enabled us to control the targeting of tumor cells as revealed by knockdown of the tumor-associated protein survivin. Overall, this study demonstrates the high potential of (networklike) streptavidin-biotin interactions with a dynamic character in the formation of complex BHS and extracellular matrix materials.

Poly(propyleneimine) glycodendrimers are proposed as nanocarriers for triphosphate forms of anticancer adenosine analogues to improve the efficiency of chemotherapy and to overcome drug resistance mechanisms. This approach has proven successful for fludarabine administration-an autonomous way of cellular entry of a nucleotide-dendrimer noncovalent complex enables an increase in the intracellular accumulation and cytotoxic activity of the active metabolite of the drug. However, the attempt to apply an analogous strategy for clofarabine results in the inhibition of drug activity. To better understand this phenomenon, characterization and comparison of drug-dendrimer complexes were needed to indicate the differences in their surface properties and the strengths of fludarabine-dendrimer and clofarabine-dendrimer interactions. Here, zeta potential measurements, ultrafiltration, and asymmetric flow field-flow fractionation are applied to determine the surface electrostatic potential and stability of nucleotide-dendrimer formulations. This approach significantly extends the authors' research on the complexation potential of perfectly branched macromolecules, ultimately explaining previously observed differences and their consequences.

Understanding the diffusion of nanoparticles through permeable membranes in cell mimics paves the way for the construction of more sophisticated synthetic protocells with control over the exchange of nanoparticles or biomacromolecules between different compartments. Nanoparticles postloading by swollen pH switchable polymersomes is investigated and nanoparticles locations at or within polymersome membrane and polymersome lumen are precisely determined. Validation of transmembrane diffusion properties is performed based on nanoparticles of different origin—gold, glycopolymer protein mimics, and the enzymes myoglobin and esterase—with dimensions between 5 and 15 nm. This process is compared with the in situ loading of nanoparticles during polymersome formation and analyzed by advanced multiple‐detector asymmetrical flow field‐flow fractionation (AF4). These experiments are supported by complementary i) release studies of protein mimics from polymersomes, ii) stability and cyclic pH switches test for in polymersome encapsulated myoglobin, and iii) cryogenic transmission electron microscopy studies on nanoparticles loaded polymersomes. Different locations (e.g., membrane and/or lumen) are identified for the uptake of each protein. The protein locations are extracted from the increasing scaling parameters and the decreasing apparent density of enzyme‐containing polymersomes as determined by AF4. Postloading demonstrates to be a valuable tool for the implementation of cell‐like functions in polymersomes.

Synthetic cells demand simultaneous and sequential tasks for developing next generation therapeutics. In article number 1801299, Albena Lederer, Dietmar Appelhans, and co‐workers report the integration of cell‐like functions in polymersomes. Through post‐loading of polymersomes, the control over protein location is discussed, essential for the exchange of biologically active materials in synthetic cells. Cover image designed by Dr. Josef Brandt.

The challenge of effective integration and use of artificial organelles with orthogonal‐responsive membranes and their communication in eukaryotic protocells is to understand the intrinsic membrane characteristics. Here, a novel photo‐crosslinked and pH‐responsive polymersome (Psome B) with 2‐(N,N′‐diisopropylamino)ethyl units in the membrane and its respective Avidin‐Psome B hybrids, are reported as good candidates for artificial organelles. Biotinylated (macro)molecules are able to dock and diffuse into Avidin‐Psome B to carry out biological activity in a pH‐ and size‐dependent manner. Combined with another polymersome (Psome A) with 2‐(N,N′‐diethylamino)ethyl units in the membrane, two different pH‐responsive polymersomes for mimicking different organelles in one protocell system are reported. The different intrinsic docking and diffusion processes of cargo (macro)molecules through the membranes of coexisting Psome A and B are pH‐dependent as confirmed using pH titration–dynamic light scattering (DLS). Psome A and B show separated “open”, “closing/opening”, and “closed” states at various pH ranges with different membrane permeability. The results pave the way for the construction of multicompartmentalized protocells with controlled communications between different artificial organelles., For studying orthogonal‐responsive membranes in eukaryotic protocells, artificial organelles are constructed by two different pH‐stable and pH‐responsive avidin–polymersome biohybrid structures. Different pH‐dependent “open”, “closing/opening”, and “closed” membrane states of the two coexisting biohybrid structures pave the way to different intrinsic and sequential docking and diffusion processes of cargo (macro)molecules in a pH‐ and size‐dependent manner.

Well-defined enzymatic biohybrid structures (BHS) composed of avidin, biotinylated poly(propyleneimine) glycodendrimers, and biotinylated horseradish peroxidase were fabricated by a sequential polyassociation reaction to adopt directed enzyme prodrug therapy to protein-glycopolymer BHS for potential biomedical applications. To tailor and gain fundamental insight into pivotal properties such as size and molar mass of these BHS, the dependence on the fabrication sequence was probed and thoroughly investigated by several complementary methods (e.g., UV/vis, DLS, cryoTEM, AF4-LS). Subsequent purification by hollow fiber filtration allowed us to obtain highly pure and well-defined BHS. Overall, by rational design and control of preparation parameters, e.g., fabrication sequence, ligand-receptor stoichiometry, and degree of biotinylation, well-defined BHS with stable and even strongly enhanced enzymatic activities can be achieved. Open coil-like structures of BHS with few branches are available by the sequential bioconjugation approach between synthetic and biological macromolecules possessing similar size dimensions.

Key parameters allow a reproducible polyassociation between avidin and biotinylated glycopolymers in order to fabricate defined supramolecular nanostructures for future (bio)medical and biotechnological applications. Thus, the polymerization efficiency of biotinylated glycopolymers in the fabrication of biohybrid structures (BHS) was investigated with regard to the influence of (i) the degree of biotinylation of the dendritic glycoarchitectures, (ii) two biotin linkers, (iii) the dendritic scaffold (perfectly branched vs hyperbranched), and (iv) the ligand-receptor stoichiometry. The adjustment of all these parameters opens the way to fabricate defined sizes of the final biohybrid structures as a multifunctional platform ready for their use in different applications. Various analytical techniques, including purification of BHS, were used to gain fundamental insights into the structural properties of the resulting protein-glycopolymer BHS. Finally, the elucidation of pivotal conformational properties of isolated BHS with defined sizes by asymmetrical flow field flow fractionation study revealed that they mainly possess spherical-/star-like properties. From this study, the fundamental knowledge can be likely transferred to other assemblies formed by molecular recognition processes (e.g., adamantane-β-cyclodextrin).

Dendritic polymers represent a class of materials for prospective drug delivery application. For that purpose we present the synthesis and characterization of hydrophilic, anionic core–shell architectures based on poly(ethylene imine) (PEI) as core molecule and polyamino acid chains (composed of glutamic acid or aspartic acid) as shell component. NCA polymerization is used for coupling polyamino acid chains to PEI scaffold. Modifying these structures with sugar molecules result in the formation of new core–shell architectures combining a mixture of binary and double shell. For their potential biomedical applications the solution properties of these anionic core–shell architectures at various pH values (3–9) were studied by different analytical tools (zeta potential, streaming potential pH titration, DLS, AFM, in-situ AFM, TEM and cryo-TEM). Especially, the sugar-decorated core–shell architectures mainly provide isolated macromolecules over a broad pH range. Furthermore, the anionic core–shell architectures are suited to interact with cationic molecules.

Aqueous solutions of multiarm flower-like poly(ethylene oxide) (PEO) were formed and connected to various degrees by self-assembly. The structure was rendered permanent by in situ UV-irradiation. Dense suspensions of these single and connected soft colloids were studied by static and dynamic light scattering and viscosity measurements. The concentration dependence of the osmotic compressibility, the dynamic correlation length, and the viscosity of single flowers was shown to be close to that of equivalent PEO star-like polymers demonstrating that the effect of forming loops on the interaction is small. It was found that the osmotic compressibility and the dynamic correlation length of dense suspensions are not influenced by the bridging. However, when flower polymers are connected into clusters, motion in dense suspensions needs to be collective over larger length scales. This causes a much stronger increase of the viscosity for dense suspensions of interpenetrated clusters compared to single-flower polymers.