Random Coil Polymers Research Articles

Ensemble structure of the N-terminal domain (1–267) of FUS in a biomolecular condensate

Solutions of some proteins phase separate into a condensed state of high protein concentration and a dispersed state of low concentration. Such behavior is observed in living cells for a number of RNA-binding proteins that feature intrinsically disordered domains. It is relevant for cell function via the formation of membraneless organelles and transcriptional condensates. On a basic level, the process can be studied in vitro on protein domains that are necessary and sufficient for liquid-liquid phase separation (LLPS). We have performed distance distribution measurements by electron paramagnetic resonance for 13 sections in an N-terminal domain (NTD) construct of the protein fused in sarcoma (FUS), consisting of the QGSY-rich domain and the RGG1 domain, in the denatured, dispersed, and condensed state. Using 10 distance distribution restraints for ensemble modeling and three such restraints for model validation, we have found that FUS NTD behaves as a random-coil polymer under good-solvent conditions in both the dispersed and condensed state. Conformation distribution in the biomolecular condensate is virtually indistinguishable from the one in an unrestrained ensemble, with the latter one being based on only residue-specific Ramachandran angle distributions. Over its whole length, FUS NTD is slightly more compact in the condensed than in the dispersed state, which is in line with the theory for random coils in good solvent proposed by de Gennes, Daoud, and Jannink. The estimated concentration in the condensate exceeds the overlap concentration resulting from this theory. The QGSY-rich domain is slightly more extended, slightly more hydrated, and has slightly higher propensity for LLPS than the RGG1 domain. Our results support previous suggestions that LLPS of FUS is driven by multiple transient nonspecific hydrogen bonding and π−sp2 interactions.

Stochastic Electrical Detection of Single Ion-Gated Semiconducting Polymers.

Semiconducting polymer chains constitute the building blocks for a wide range of electronic materials and devices. However, most of their electrical characteristics at the single-molecule level have received little attention. Elucidating these properties can help understanding performance limits and enable new applications. Here, coupled ionic-electronic charge transport is exploited to measure the quasi-1D electrical current through long single conjugated polymer chains as they form transient contacts with electrodes separated by ≈10nm. Fluctuations between internal conformations of the individual polymers are resolved as abrupt, multilevel switches in the electrical current. This behavior is consistent with the theoretical simulations based on the worm-like-chain (WLC) model for semiflexible polymers. In addition to probing the intrinsic properties of single semiconducting polymer chains, the results provide an unprecedented window into the dynamics of random-coil polymers and enable the use of semiconducting polymers as electrical labels for single-molecule (bio)sensing assays.

PEG-Lipids: Quantitative Study of Unimers and Aggregates Thereof by the Methods of Molecular Hydrodynamics.

Understanding the polymorphism of lipids in solution is the key to the development of intracellular delivery systems. Here, we study the dynamics of poly(ethylene glycol)-lipid (PEG-Lipid) conjugates aiming at a better understanding of their molecular properties and aggregation behavior in solution. Those PEG-Lipids are used as components of lipid nanoparticles (LNPs). LNPs are gaining increased popularity, e.g., by their utilization in modern vaccination strategies against SARS-CoV-2. Characterization of the systems is conducted by the classical methods of hydrodynamics in different solvents, such as ethanol and water, which are also commonly used for LNP formulation. We were able to elucidate the structurally associated hydrodynamic properties of isolated PEG-Lipids in ethanol, revealing the typically expected values of the hydrodynamic invariant for random coil polymers. By virtue of the same experimental setting, the PEG-Lipids' behavior in water was as well studied, which is a less good solvent than ethanol for the PEG-Lipids. Our experiments demonstrate that PEG-Lipids dissolved in water form well-defined micelles that can quantitatively be characterized in terms of their degree of aggregation of PEG-Lipid polymer unimers, their hydrodynamic size, and solvation, i.e., the quantitative determination of water contained or associated to the identified micelles. Quantitative results obtained from classical hydrodynamic analyses are fully supported by studies with standard dynamic light scattering (DLS). The obtained diffusion coefficients and hydrodynamic sizes are in excellent agreement with numerical results derived from analytical ultracentrifugation (AUC) data. Cryo-transmission electron microscopy (cryo-TEM) supports the structural insight from hydrodynamic studies, particularly, in terms of the observed spherical structure of the formed micelles. We demonstrate experimentally that the micelle systems can be considered as solvent-permeable, hydrated spheres.

Secondary Structure in Overcoming Photosensitizers' Aggregation: α-Helical Polypeptides for Enhanced Photodynamic Therapy.

Aggregation caused quenching (ACQ) effect could severely inhibit the application of hydrophobic photosensitizers (PSs) bearing planar and rigid structures. Most of the reported cases utilized random-coiled polymers for the in vivo delivery of PSs, which would inevitably aggravate ACQ effect due to the flexible chains. In this work, the role of polymers' secondary structures (especially α-helical conformation) in overcoming the PSs' aggregation was systemically investigated based on the design of α-helical polypeptides bearing tetraphenylporphyrin (TPP) side chains. Atomistic molecular dynamics simulation, fluorescence quantum yield and reactive oxygen species (ROS) generation yield were evaluated to demonstrate that α-helical polypeptide backbones could significantly boost both fluorescence quantum yield and ROS by suppressing the π-π stacking interaction between TPP units. The enhanced in vitro and in vivo phototoxicity for helical polypeptides also revealed functions of secondary structures in inhibiting ACQ and improving the membrane activity. Successful in vivo photodynamic therapy (PDT) results in mice bearing H22 tumors showed great potentials for further clinical applications. This article is protected by copyright. All rights reserved.

Defect-Free Synthesis of a Fully π-Conjugated Helical Ladder Polymer and Resolution into a Pair of Enantiomeric Helical Ladders.

Fully π-conjugated ladder polymers with a spiral geometry represent a new class of helical polymers with great potential for organic nanodevices, but there is no precedent for an optically-active helical ladder polymer totally composed of achiral units. We now report the defect-free synthesis and resolution of a fully π-conjugated helical ladder polymer with a rigid helical cavity, which has been achieved by quantitative and chemoselective acid-promoted alkyne benzannulations of a rationally designed, random-coil achiral polymer followed by chromatographic enantioseparation. Because of a sufficiently high helix-inversion barrier, the isolated excess one-handed helical ladder polymer with the degree of polymerization of more than 15 showed a strong circular dichroism with the dissymmetry factor of up to 1.7 × 10-2 and is thermally stable, maintaining its optical activity in solution even at 100 °C, as well supported by molecular dynamics simulation.

One-Handed Helical Tubular Ladder Polymers for Chromatographic Enantioseparation.

Defect-free one-handed contracted helical tubular ladder polymers with a π-electron-rich cylindrical helical cavity were synthesized by alkyne benzannulations of the random-coil precursor polymers containing 6,6'-linked-1,1'-spirobiindane-7,7'-diol-based chiral monomer units. The resulting tightly-twisted helical tubular ladder polymers showed remarkably high enantioseparation abilities toward a variety of chiral hydrophobic aromatics with point, axial, and planar chiralities. The random-coil precursor polymer and analogous rigid-rod extended helical ribbon-like ladder polymer with no internal helical cavity exhibited no resolution abilities. The molecular dynamics simulations suggested that the π-electron-rich cylindrical helical cavity formed in the tightly-twisted tubular helical ladder structures is of key importance for producing the highly-enantioseparation ability, by which chiral aromatics can be enantioselectively encapsulated by specific π-π and/or hydrophobic interactions.

A Sulfonated All-Aromatic Polyamide for Heavy Metal Capture: A Model Study with Pb(II).

Polyelectrolytes are widely used in heavy metal removal, finding applications as coagulants and flocculants. We compare the heavy metal removal capability of a water-soluble sulfonated semirigid polyamide, poly(2,2'-disulfonyl-4,4'-benzidine isophthalamide) (PBDI), with that of a well-known random-coil polymer, poly(sodium 4-styrenesulfonate) (PSS). Using lead (Pb(II)) as a model contaminant, both polymers precipitate out from solution at ~500 mg/L Pb(II) in water. The ability to remove Pb(II) from water was quantified using adsorption isotherms and fitted with Langmuir and Freundlich adsorption models. The sorption of Pb(II) by PSS fit the Langmuir model with a high degree of correlation (0.976 R2), but the sorption of Pb(II) by PBDI could not be accurately predicted using the Langmuir or Freundlich model. The sorption of Pb(II) by PBDI and PSS was compared by normalizing sorption by the number of sulfonate groups of each polymer and the ion exchange capacity (IEC), found by titration. We find that PBDI removes a greater amount of Pb(II) per gram of sorbent compared to PSS, 410 mg/g vs 260 mg/g, respectively, which cannot be accounted for by differences in IEC or number of sulfonate groups. Our findings confirm that the positioning of the sulfonate groups and the rigidity of the polymer backbone play an important role in how Pb(II) coordinates to the polymer prior to precipitating out from solution.

Enhanced Host-Guest Association and Fluorescence in Copolymers from Copper Salphen Complexes by Supramolecular Internalization of Anions.

We describe the synthesis, crystallographic characterization of a new Cu-Salphen compound and its use as a host Lewis-acid against guest anions in two versions: a) free molecule, b) copolymerized with methyl methacrylate:n-butyl acrylate (1 : 4-wt.) as protective co-monomers. Higher contents in Cu-Salphen yielded larger and more homogeneous polymer sizes. Polymer size together with glass transitions, heat capacity, thermal degradation, guest-saturation degrees and host-guest species distribution profiles from spectrophotometric titrations explained growths of up to 630-fold in K11 and 180000-fold in K12 for the host's binding site attributable to a solvophobic protection from the macromolecular structure. Spectrofluorimetry revealed blue-shifted×13-16 larger luminescence for Cu-Salphen in the polymers (λem =488-498 nm) than that of the non-polymerized counterpart (λem =510-543 nm) and "turn-on" blue-shifted enhanced fluorescence upon guest association. We propose a cooperative incorporation of the guests occurring from the outer medium toward internally protected binding site pockets in the random coil polymer conformations.

Selective Transport in the Nuclear Pore Complex

Nuclear pore complexes (NPCs) mediate nucleocytoplasmic exchange controlling the flow of molecules into and out of the nucleus. The selective filter properties of NPCs enable translocation of specific molecules known as nuclear transport factors (NTRs) and their cargo. Central to this selectivity barrier is a group of largely intrinsically disordered nucleoporins (Nups) that contain multiple phenylalanine‐glycine repeats, termed FG Nups. The interactions between FG Nups and NTRs enable NTRs to translocate rapidly yet selectively through the NPC in what is referred to as the “transport paradox”. FG Nups in the NPC have a small folded anchor region tethering them to the NPC outer ring and otherwise are fully disordered, random coil polymers that remain predominantly disordered while engaged to NTRs. FG Nups interact with NTRs using mainly their FG motifs and minimally their intervening spacer residues. The overall enthalpy of the interaction increases as multivalency increases the frequency of individually weak FG‐NTR contacts. Tight binding is limited by an entropy penalty that disfavors simultaneous engagements of FG motifs. Small angle neutron scattering (SANS) shows that the entropy loss is partly due to the local rigidity of an FG motif in the interacting state(Fig 1). All‐atom molecular dynamics (MD) simulation indicates that spacers between the FG motifs behave as “entropic springs”, disfavoring any static binding of the FG repeats. The dynamics of FG Nups enables the FG motifs to slide along the hydrophobic patches of NTRs enabling FG motifs to be easily displaced by other competing FG motifs (Fig. 2). This explanation provides a simple hypothesis for the rapid exchange of FG motif contacts during transport, focusing on the entropic exclusion of non‐NTRs, and ‘solubilization’ of NTR complexes, in contrast to possible condensate formation which would provide a compartmentalization of components. These results reveal fundamental aspects of the functioning mechanisms underlying NPC transport at high structural resolution, something lacking in current models of nuclear transport.1. Sparks, S., et al., Analysis of Multivalent IDP Interactions: Stoichiometry, Affinity, and Local Concentration Effect Measurements. Methods Mol Biol, 2020. 2141: p. 463‐475.2. Sparks, S., et al., Deciphering the “Fuzzy” Interaction of FG Nucleoporins and Transport Factors Using Small‐Angle Neutron Scattering. Structure, 2018. 26(3): p. 477‐484 e4.3. Hayama, R., et al., Thermodynamic characterization of the multivalent interactions underlying rapid and selective translocation through the nuclear pore complex. J Biol Chem, 2018. 293(12): p. 4555‐4563.4. Hough, L.E., et al., The molecular mechanism of nuclear transport revealed by atomic‐scale measurements. Elife, 2015. 4.5. Raveh, B., et al., Slide‐and‐exchange mechanism for rapid and selective transport through the nuclear pore complex. Proc Natl Acad Sci U S A, 2016. 113(18): p. E2489‐97.

Supersoft Elastic Bottlebrush Microspheres with Stimuli-Responsive Color-Changing Properties in Brine.

Solvent-free supersoft elastomer is highly desirable for building photonic structures with significant stimuli-responsive color changes. We report supersoft elastic porous microspheres with vivid structural colors obtained via self-assembly of amphiphilic bottlebrush block copolymers at the water/oil interface templated by ordered water-in-oil-in-water double emulsions. The porous structure is composed of cross-linked bottlebrush polydimethylsiloxane (PDMS) as the supersoft elastic skeleton and bottlebrush poly(ethylene oxide) (PEO) as the internal responsive layer. The obtained microspheres show large reversible volume changes through well-controlled dehydration or hydration of PEO in response to salt ions in an aqueous environment. As a result, full-spectrum colors are obtained dependent on different salt concentrations. In-situ observation of color reflection of a microsphere indicates a gradual structural transition from the outside to the inside corresponding to migration of water molecules and salt ions. Moreover, rod-like bottlebrush PEO exhibits an anion-induced salting-out behavior different from that of random coil polymers. The significantly responsive behaviors of bottlebrush block copolymer (BBCP) assemblies in the presence of salt ions primarily rely on the supersoft elastic skeleton of the porous structure, providing a facile route to the creation of stimuli-responsive photonic materials by low-cost self-assembly methods.

Amino-acid-substituted polyacetylene-based chiral core–shell microspheres: helix structure induction and application for chiral resolution and adsorption

The random coil polymer was first compounded with substrate and induced it into helical structure subsequently.

Polymer Stiffness Regulates Multivalent Binding and Liquid-Liquid Phase Separation

Multivalent binding is essential to many biological processes because it builds high-affinity bonds by using several weak binding interactions simultaneously. Multivalent polymers have shown promise as inhibitors of toxins and other pathogens, and they are important components in the formation of biocondensates. Explaining how structural features of these polymers change their binding and subsequent control of phase separation is critical to designing better pathogen inhibitors and also to understanding diseases associated with membraneless organelles. In this work, we will examine the binding of a multivalent polymer to a small target. This scenario could represent a polymeric inhibitor binding to a toxic protein or RNA binding to an RNA-binding protein in the case of liquid-liquid phase separation. We use simulation and theory to show that flexible random-coil polymers bind more strongly than stiff rod-like polymers and that flexible polymers nucleate condensed phases at lower binding energies than their rigid analogs. We hope these results will provide insight into the rational design of polymeric inhibitors and improve our understanding of phase separation in cells and membraneless organelles.

Stretchable hydrogels with low hysteresis and anti-fatigue fracture based on polyprotein cross-linkers

Hydrogel-based devices are widely used as flexible electronics, biosensors, soft robots, and intelligent human-machine interfaces. In these applications, high stretchability, low hysteresis, and anti-fatigue fracture are essential but can be rarely met in the same hydrogels simultaneously. Here, we demonstrate a hydrogel design using tandem-repeat proteins as the cross-linkers and random coiled polymers as the percolating network. Such a design allows the polyprotein cross-linkers only to experience considerable forces at the fracture zone and unfold to prevent crack propagation. Thus, we are able to decouple the hysteresis-toughness correlation and create hydrogels of high stretchability (~1100%), low hysteresis (< 5%), and high fracture toughness (~900 J m−2). Moreover, the hydrogels show a high fatigue threshold of ~126 J m−2 and can undergo 5000 load-unload cycles up to 500% strain without noticeable mechanical changes. Our study provides a general route to decouple network elasticity and local mechanical response in synthetic hydrogels.

Macromolecular Chiral Amplification through a Random Coil to One-Handed Helix Transformation Induced by Metal Ion Coordination in an Aqueous Solution

Many inspiring achievements have been made in inducing chiral amplification in helical polymers; however, the induction of helical chirality in random coil polymers by the intermolecular noncovalen...

Conformation of Tunable Nanocylinders: Up to Sixth-Generation Dendronized Polymers via Graft-Through Approach by ROMP.

Well-defined dendronized polymers (denpols) bearing high-generation dendron are attractive nano-objects as high persistency provides distinct properties, contrast to the random coiled linear polymers However, their syntheses via graft-through approach have been very challenging due to their structural complexity and steric hindrance retarding polymerization. Here, we report the first example of the synthesis of poly(norbornene) (PNB) containing ester dendrons up to the sixth generation (G6) by ring-opening metathesis polymerization. This is the highest generation ever polymerized among dendronized polymers prepared by graft-through approach, producing denpols with molecular weight up to 1960 kg/mol. Combination of size-exclusion chromatography, light scattering, and neutron scattering allowed a thorough structural study of these large denpols in dilute solution. A semiflexible cylinder model was successfully applied to represent both the static and dynamic experimental quantities yielding persistent length (lp), cross-sectional radius (Rcs), and contour length (L). The denpol persistency seemed to increase with generation, with lp reaching 27 nm (Kuhn length 54 nm) for PNB-G6, demonstrating a rod-like conformation. Poly(endo-tricycle[4.2.2.0]deca-3,9-diene) (PTD) denpols exhibited larger persistency than the PNB analogues of the same generation presumably due to the higher grafting density of the PTD denpols. As the dendritic side chains introduce shape anisotropy into the denpol backbone, future work will entail a study of these systems in the concentrated solutions and melts.

POLYMER APPARENT VISCOSITY DEPENDENCE ON INACCESSIBLE PORE VOLUME: LABORATORY AND FIELD STUDIES OF ITS INFLUENCE ON ENHANCED OIL RECOVERY

The successful use of polymer-enhanced oil recovery observed in the last two decades is leading to effective field implementations. One of the reasons for such positive results in polymer flooding is the integration between laboratory-measured properties and reservoir simulation. Recent reports show that inaccessible pore volume (IAPV) plays a significant role in the apparent viscosity of random coil polymers, such as hydrolyzed polyacrylamide (HPAM). This paper assesses both direct and indirect impacts of IAPV on polymer flooding. This investigation relies on laboratory and field-scale simulations of polymer flooding using CMG-STARS. Simulation cases review the effects of IAPV on production indicators in direct and indirect forms. This study compares laboratory-scale simulations with experimental results and uses quality indicators to evaluate the history matching of both approaches. It relies on a modified benchmark field case to study both approaches in comparison to waterflooding and idealized polymer flooding. Results indicate that IAPV has a small direct impact on production curves. However, data show significant indirect implications of the IAPV on recovery. This effect occurs because the apparent viscosity of HPAM has a direct relationship with IAPV. Although the simulation results were consistent with current literature, the results obtained through the experiments indicate that the most realistic simulation case is achieved when considering the impact of IAPV on polymer viscosity.

Fabrication of In Situ Nanofiber-Reinforced Molecular Composites by Nonequilibrium Self-Assembly.

Although the concept of molecular composites (MCs) is very promising, there are major obstacles arising from the immiscibility of the rigid-rod with the random-coil polymers. Here, we developed a novel method for fabricating an in situ reinforced MC system with nonequilibrium self-assembled nanofibrous structures based on bisphenol A epoxy resin, 4,4'-diaminodiphenylsulfone, bismaleimide, and a polyphenylene ether (PPO) oligomer. A variety of spectroscopic and morphological techniques were used to probe the structural evolution from the emergence of nanofibrils, to growth and aggregation of nanofibers, and then to the formation of in situ reinforced MC with strong interfacial interactions. The in situ nanofibers within the polymer matrix could be formed by the polymerization force extruding the PPO phase through the interspaces within the simultaneous interpenetrating network polymers during the cure process of the thermosetting resin system. Compared to the control sample, the in situ nanofiber-reinforced MC exhibited better thermal properties and flame retardancy. In particular, the obtained MC showed a significant improvement in glass transition temperature and mechanical properties, which were mainly attributed to the restriction of high thermal stability of PPO on the segmental motion of polymer chains, the toughening and reinforcement behaviors of PPO nanofibers on the matrix, and the chemical interaction at the PPO/matrix interface.

DNA-Based Nanodevices Controlled by Purely Entropic Linker Domains.

We demonstrate here the rational design of purely entropic domains as a versatile approach to achieve control of the input/output response of synthetic molecular receptors. To do so and to highlight the versatility and generality of this approach, we have rationally re-engineered two model DNA-based receptors: a clamp-like DNA-based switch that recognizes a specific DNA sequence and an ATP-binding aptamer. We show that, by varying the length of the linker domain that connects the two recognition portions of these receptors, it is possible to finely control their affinity for their specific ligand. Through mathematical modeling and thermodynamic characterization, we also demonstrate for both systems that entropy changes associated with changes in linker length are responsible for affinity modulation and that the linker we have designed behaves as a disordered random-coil polymer. The approach also allows us to regulate the ligand concentration range at which the receptors respond and show optimal specificity. Given these attributes, the use of purely entropic domains appears as a versatile and general approach to finely control the activity of synthetic receptors in a highly predictable and controlled fashion.

Ultrafast and Efficient Detection of Formaldehyde in Aqueous Solutions Using Chitosan-based Fluorescent Polymers.

Detection of a toxic formaldehyde (HCHO) pollutant in aqueous solutions is of significant importance, because HCHO is widely found in aquatic food because of illicit addition or improper storage. Many small-molecule-based fluorescent probes, which rely on HCHO-specific formaldehyde-amine condensation or the aza-Cope rearrangement reaction, have been developed in terms of facile operation and high selectivity. However, some primary challenging issues are the restricted sensitivity and long equilibrium response time caused by the slow chemical reaction between these small-molecule-based sensors and low-concentration HCHO pollutant in testing samples. Herein, a robust hydrophilic hydrazino-naphthalimide-functionalized chitosan (HN-Chitosan)-based polymeric probe is reported, which takes advantage of specific chemical reaction between HCHO and grafted hydrazino-naphthalimide groups to trigger a "turn-on" fluorescence response. Superior to its small-molecule analogs, HN-Chitosan is based on random coil polymer chains of biopolymeric chitosan, which is thus capable of employing the cooperative binding effect of multiple hydrazino-naphthalimide recognition sites and adjacent hydroxyl groups to "enrich" the low-concentration HCHO pollutant around the polymer chains via weak supramolecular interactions. Therefore, the HCHO-specific chemical reaction with grafted hydrazino-naphthalimide groups is significantly accelerated, resulting in the unprecedented ultrafast equilibrium fluorescence response (less than 1 min) and high sensitivity. Encouraged by its satisfying sensitivity, selectivity, fast response, and wide linear detection range, we successfully expand its application to real-world food and water analysis. In view of the modular design principle of our polymeric probe, the proposed strategy could be generally applicable to construct powerful polymeric probes for ultrafast detection of other important pollutants.

Excluded Volume Effects Inside the Living Cell

Proteins and nucleic acids fold and behave in a highly occupied matrix of macromolecules, the cellular environment. Although this environment is filled up to a volume of 40% with macromolecules, the effect of crowding on biochemical reactions as well as biophysical properties of proteins has been rarely considered. Therefore, studies on macromolecular crowding are mainly conducted using artificially created polymer based substances as mimics of the in cell environment. Still, a common concept how the excluded volume effect in the cell affects protein folding, function and structure is lacking. Understanding these properties will be important to unravel the underlying mechanism of protein misfolding and aggregation as well as the behavior of intrinsically disordered proteins in a living cell.We introduce a FRET based random coil polymer to characterize crowding both in vitro and inside a living cell. We find different compression of the polymer in artificial crowding substances compared to protein crowders such as BSA or oocyte lysate. Injection of the probe inside cells confirms this result and reveals a heterogeneous environment which, on average, shows comparable polymer conformations as in diluted buffer. The polymer conformation is used to quantify crowding differences in the cytosol and the nucleus as well as to identify crowding in cells influenced by different extrinsic conditions. Severe osmotic stress is used to probe compression of the polymer in the highly concentrated cytosolic environment.We conclude that the FRET labelled polymer provides a new approach to investigate and characterize the cellular solvation properties with high spatio-temporal resolution in a variety of systems and identify crowding differences due to the architecture of the cellular matrix. Therefore, it will help to understand how the cell, as the native environment in which proteins evolved, might modulate and tune biomolecule properties and functions.