Effect of Physical Properties of Nanogel Particles on the Kinetic Constants of Multipoint Protein Recognition Process

We report the effect of physical properties, such as flexibility and polymer density, of nanogel particles (NPs) on the association/dissociation rates constant (<i>k</i>_on and <i>k</i>_off) and equilibrium constants (<i>K</i>_d) of multipoint protein recognition process. NPs having different flexibilities and densities at 25 °C were synthesized by tuning cross-linking degrees and the volume phase transition (VPT) temperature. Rate constants were quantified by analyzing time course of protein binding process on NPs monitored by a quartz crystal microbalance (QCM). Both <i>k</i>_on and <i>k</i>_off of swollen phase NPs increased with decreasing cross-linking degree, whereas cross-linking degree did not affect <i>k</i>_on and <i>k</i>_off of the collapsed phase NPs, indicating that polymer density of NPs governs <i>k</i>_on and <i>k</i>_off. The results also suggest that the mechanical flexibility of NPs, defined as the Young’s modulus, does not always have crucial roles in the multipoint molecular recognition process. On the other hand, <i>K</i>_d was independent of the cross-linking degree and depended only on the phase of NPs, indicating that molecular-scale flexibility, such as side-chain and segmental-mode mobility, as well as the conformation change, of polymer chains assist the formation of stable binding sites in NPs. Our results reveal the rationale for designing NPs having desired affinity and binding kinetics to target molecules

Control of Protein-Binding Kinetics on Synthetic Polymer Nanoparticles by Tuning Flexibility and Inducing Conformation Changes of Polymer Chains

Although a number of procedures to create synthetic polymer nanoparticles (NPs) with an intrinsic affinity to target biomacromolecules have been published, little has been reported on strategies to control the binding kinetics of target recognition. Here, we report an enzyme-mimic strategy to control binding/dissociation rate constants of NPs, which bind proteins through multipoint interactions, by taking advantage of the temperature-responsive coil–globule phase transition of poly-<i>N</i>-isopropylacrylamide (PNIPAm)-based NPs. PNIPAm NPs with a “flexible” random-coil conformation had a faster binding rate than NPs with a “rigid” globule conformation; however, the dissociation rate constant remained unchanged, resulting in stronger affinity. The dissociation rate of the “flexible” NPs was decelerated by the “induced-fit”-type conformation change of polymers around the coil–globule phase transition temperature, resulting in the formation of the most stable NP–protein complexes. These results provide a guide for designing plastic antibodies with tailor-made binding kinetics and equilibrium constants

Self-Assembly of a Double Hydrophilic Block Glycopolymer and the Investigation of Its Mechanism

We report the self-assembly of a double hydrophilic block glycopolymer (DHBG) via hydrogen bonding and coordinate bonding. This DHBG, composed of poly­(ethylene)­glycol (PEG) and glycopolymer, self-assembled into a well-defined structure. The DHBG was prepared through the controlled radical polymerization of trimethylsilyl-protected propargyl methacrylate using a PEG-based reversible addition–fragmentation chain transfer reagent, followed by sugar conjugation using click chemistry. The DHBG self-assembly capability was investigated by transmission electron microscopy and dynamic light scattering. Interestingly, the DHBG self-assembled into a spherical structure in aqueous solution. Hydrogen bonding and coordinate bonding with Ca²⁺ were identified as the driving forces for self-assembly

Design of Synthetic Polymer Nanoparticles That Facilitate Resolubilization and Refolding of Aggregated Positively Charged Lysozyme

Designed polymer hydrogel nanoparticles (NPs) capable of facilitating resolubilization and refolding of an aggregated protein, positively charged lysozyme, are prepared. NPs designed to interact strongly with denatured lysozyme and relatively weakly with native lysozyme, facilitated resolubilization and refolding of aggregated lysozyme. Such NPs could be prepared by copolymerizing optimized combinations and populations of functional monomers. The refolded lysozyme showed native conformation and enzymatic activity. Eleven grams of aggregated protein was refolded by 1 g of NPs. However, NPs having low affinity to denatured lysozyme and NPs having high affinity to both denatured and native lysozyme showed relatively low facilitation activity. Our results suggest a potential strategy for the design of artificial chaperones with high facilitating activity

Reversible Absorption of CO₂ Triggered by Phase Transition of Amine-Containing Micro- and Nanogel Particles

Herein we report that an aqueous solution of temperature-responsive micro- and nanogel particles (GPs) consisting of <i>N</i>-isopropylacrylamide (NIPAm) and <i>N</i>-[3-(dimethylamino)­propyl]­methacrylamide (DMAPM) reversibly absorbs and desorbs CO₂ via a phase transition induced by cooling and heating cycles (30–75 °C). Below the phase-transition temperature, most of the amines in the swollen GPs are capable of forming ion pairs with absorbed bicarbonate ions. However, above the phase-transition temperature, shrinkage of the GPs lowers the p<i>K</i>_a and the number of amine groups exposed to water, thereby resulting in almost complete desorption of CO₂. The GPs can reversibly absorb more than the DMAPM monomer and polymer without NIPAm, which indicates the importance of the temperature-responsive phase transition of polymers in determining the degree of absorption. The results show the potential of temperature-responsive polymer solutions as absorbents to sequester CO₂ at a low energy cost

Optimization of Poly(<i>N</i>‑isopropylacrylamide) as an Artificial Amidase

Poly­(<i>N</i>-isopropylacrylamide) microgel (NMG) has been developed by adding various functional groups to control surface charges, hydrophobicity, p<i>K</i>_a and protein adsorption capacity. Here, we developed and optimized NMG anchored with three types of functional groups as a polymeric catalyst to hydrolyze amide bonds under optimized mild conditions. Various optimization strategies were evaluated for efficient hydrolysis activity on a <i>p</i>-nitroaniline-based substrate by using a colorimetric assay. Based on the results, we propose a mechanism to hydrolyze amide bonds and determine the theoretical average distance, using NMG bearing functional group of 1-vinylimidazole as the study model. The hydrolysis of amide bonds was inhibited by a transition-state protease inhibitor, which also confirmed the proposed reaction model for NMG. These results provide an insight into the strategies developed to functionalize hydrogels through an enzyme-mimic approach for future robust bio- and chemical conversions as well as therapeutic utilities

Macroporous Gel with a Permeable Reaction Platform for Catalytic Flow Synthesis

We mimic a living system wherein target molecules permeate through capillary and cells for chemical transformation. A monolithic porous gel (MPG) was easily prepared by copolymerization of gel matrix, tertiary amine, and cross-linking monomer in one-step synthesis. Interconnected capillaries existed in the MPG, enabling flow application with high permeability. Because the capillaries were constituted of polymer gel, Pd(0)-loaded MPG provided another permeable pathway to substrates in a gel network, contributing to its much high turnover number after 30 days of use, compared with that of Pd(0)-loaded inorganic supports. Interestingly, the gel network size of the MPG influenced the catalytic frequency. Diffusivities of the substrates and product in the gel networks increased with increasing network sizes in relation to catalytic activities. The MPG strategy provides a universal reactor design in conjunction with a practical process and precisely controlled reaction platform

Design of Glycopolymers Carrying Sialyl Oligosaccharides for Controlling the Interaction with the Influenza Virus

We designed glycopolymers carrying sialyl oligosaccharides by “post-click” chemistry and evaluated the interaction with the influenza virus. The glycopolymer structures were synthesized in a well-controlled manner by reversible addition–fragmentation chain transfer polymerization and the Huisgen reaction. Acrylamide-type monomers were copolymerized to give hydrophilicity to the polymer backbones, and the hydrophilicity enabled the successful introduction of the oligosaccharides into the polymer backbones. The glycopolymers with different sugar densities and polymer lengths were designed for the interaction with hemagglutinin on the virus surface. The synthesized glycopolymers showed the specific molecular recognition against different types of influenza viruses depending on the sugar units (6′- or 3′-sialyllactose). The sugar density and the polymer length of the glycopolymers affected the interaction with the influenza virus. Inhibitory activity of the glycopolymer against virus infection was demonstrated

Inhibition of Bacterial Adhesion on Hydroxyapatite Model Teeth by Surface Modification with PEGMA-Phosmer Copolymers

Modification of the interface properties on hydroxyapatite and tooth enamel surfaces was investigated to fabricate bacterial resistance <i>in situ</i>. A series of copolymers containing pendants of poly­(ethylene glycol) methyl ether methacrylate (PEGMA) and ethylene glycol methacrylate phosphate (Phosmer) were polymerized by conventional free radical polymerization and changing the feed ratio of monomers. The copolymers were immobilized on hydroxyapatite and tooth enamel via the affinity of phosphate groups to hydroxyapatite to form the stable and durable polymer brushes on the surfaces. The amounts of polymer immobilized depended on the phosphate group ratio in the copolymers. Surface modification altered the interfacial properties of hydroxyapatite and inhibited bacterial adhesion. Copolymers containing 40–60% PEGMA segments showed a significant inhibitory effect on bacterial adhesion of <i>S. epidermidis</i> both in the presence and absence of plaque model biomacromolecules

Minimization of Synthetic Polymer Ligands for Specific Recognition and Neutralization of a Toxic Peptide

Synthetic polymer ligands (PLs) that recognize and neutralize specific biomacromolecules have attracted attention as stable substitutes for ligands such as antibodies and aptamers. PLs have been reported to strongly interact with target proteins and can be prepared by optimizing the combination and relative proportion of functional groups, by molecular imprinting polymerization, and/or by affinity purification. However, little has been reported about a strategy to prepare PLs capable of specifically recognizing a peptide from a group of targets with similar molecular weight and amino acid composition. In this study, we show that such PLs can be prepared by minimization of molecular weight and density of functional units. The resulting PLs recognize the target toxin exclusively and with 100-fold stronger affinity from a mixture of similar toxins. The target toxin is neutralized as a result. We believe that the minimization approach will become a valuable tool to prepare “plastic aptamers” with strong affinity for specific target peptides