Thiol-Reactive Clickable Cryogels: Importance of Macroporosity and Linkers on Biomolecular Immobilization

Abstract

Macroporous cryogels that are amenable to facile functionalization are attractive platforms for biomolecular immobilization, a vital step for fabrication of scaffolds necessary for areas like tissue engineering and diagnostic sensing. In this work, thiol-reactive porous cryogels are obtained via photopolymerization of a furanprotected maleimide-containing poly(ethylene glycol) (PEG)-based methacrylate (PEGFuMaMA) monomer. A series of cryogels are prepared using varying amounts of the masked hydrophilic PEGFuMaMA monomer, along with poly(ethylene glycol) methyl ether methacrylate and poly(ethylene glycol) dimethacrylate, a hydrophilic monomer and cross-linker, respectively, in the presence of a photoinitiator. Subsequent activation to the thiol-reactive form of the furan-protected maleimide groups is performed through the retro Diels−Alder reaction. As a demonstration of direct protein immobilization, bovine serum albumin is immobilized onto the cryogels. Furthermore, ligand-directed immobilization of proteins is achieved by first attaching mannose− or biotin−thiol onto the maleimide-containing platforms, followed by ligand-directed immobilization of concanavalin A or streptavidin, respectively. Additionally, we demonstrate that the extent of immobilized proteins can be controlled by varying the amount of thiol-reactive maleimide groups present in the cryogel matrix. Compared to traditional hydrogels, cryogels demonstrate enhanced protein immobilization/detection. Additionally, it is concluded that utilization of a longer linker, distancing the thiol-reactive maleimide group from the gel scaffold, considerably increases protein immobilization. It can be envisioned that the facile fabrication, conjugation, and control over the extent of functionalization of these cryogels will make these materials desirable scaffolds for numerous biomedical applications.

Succinimidyl Carbonate-Based Amine-Reactive Polymer Brushes: Facile Fabrication of Functional Interfaces

“…Furthermore, the bulk of the protein may also pose significant steric hindrance for attachment with the amine reactive group close to the polymer backbone, while a biotin group appended through a linker may facilitate access to the protein binding site because using a longer linker is known to provide better conjugation. 48,49 The strong biotin−streptavidin interaction was further used to direct nanoparticle assembly onto the SCEMA-based copolymer brushes. To this end, Streptavidin-coated Q-Dots were immobilized on biotinylated patterned copolymer brushes (P2−P4), which were analyzed with fluorescence microscopy (Figure 8).…”

“…Also, the DEGMEMA monomer is structurally related to 2-hydroxyl ethyl methacrylate and poly­(ethylene glycol) methacrylate, which are used to produce polymer brushes that are very effective in preventing nonspecific protein adsorption, thus hindering the reaction of streptavidin as compared to the amine-functional biotin molecule. Furthermore, the bulk of the protein may also pose significant steric hindrance for attachment with the amine reactive group close to the polymer backbone, while a biotin group appended through a linker may facilitate access to the protein binding site because using a longer linker is known to provide better conjugation. , …”

Nanoscale Structures of Poly(oligo ethylene glycol methyl ether methacrylate) Hydrogels Revealed by Small-Angle Neutron Scattering

“…Thus, the volume transition temperature of the copolymerized gels can be controlled from 20 to 90 °C by adjusting the copolymerization ratio between MeO 2 MA and OEGMA during the polymerization. − Moreover, POEGMA chains have many advantages, similar to those of polyethylene glycol chains, such as their nonimmunogenic, noncytotoxic, and protein- (and consequently cell)-repellent properties . Indeed, many studies related to POEGMA-based materials involve their use in advanced biomedical applications, such as bioconjugates, , surface coatings, , particles for carriers, − and injectable gels. − …”

Tailoring Aqueous Dispersibility and Biofunctionalization of Carbon Nanotubes Using Maleimide-Containing Clickable Polymers

“…Among the various possible clickable groups, such as azides, alkynes, and alkenes, functional groups have been extensively utilized in post-polymerization functionalization. In particular, the electron-deficient maleimide group has been employed as an efficient handle for the conjugation of thiol-containing (bio)­molecules for the past several decades. − Although it is not possible to directly incorporate the maleimide group during polymerization, a Diels–Alder/retro-Diels–Alder reaction-based manipulation is well established to provide facile access to the maleimide group containing polymeric materials. ,− Thus, the fabrication of CNTs coated with PEG-based hydrophilic copolymers containing the thiol-reactive maleimide groups as side chains will provide a system amenable for reagent-free functionalization to engineer the desired functional interfaces.…”