Rational Design of a Tetrameric Protein to Enhance Interactions between Self‐Assembled Fibers Gives Molecular Hydrogels

Abstract

Molecular hydrogels have attracted extensive research interest in recent years because of their inherent properties (e.g., formation by the self-assembly of small molecules and their gel-sol/sol-gel phase transitions can be easily manipulated by external stimulus). [1] They have shown great potential in fields such as three-dimensional (3D) cell culture [2] and controlled drug delivery. [3] During the formation of a molecular hydrogel, a small molecule (molecular hydrogelator) needs to selfassemble into a 3D matrix of nanofibers, nanorods, or nanospheres that can hold water molecules within the cavities of the 3D matrix. To form the 3D matrix, there should be strong or at least medium interactions between self-assembled nanostructures. Otherwise, nanostructures with weak interactions between them will only form dispersions or solutions in the aqueous phase. Actually, there are many examples of this kind of self-assembled system that lack strong interactions between the self-assembled structures. [4] This type of solution/dispersion containing self-assembled nanostructures could change to a hydrogel if the interaction between the nanostructures could be enhanced. For example, several groups have demonstrated that zinc and calcium ions can be used to cross-link self-assembled nanofibers to form molecular hydrogels. [5] In this study, we rationally designed a fusion protein with four binding sites and used the proteinpeptide interaction to enhance interactions between selfassembled nanofibers, thus leading to the formation of molecular hydrogels (Figure 1).There are only a few examples of polymeric hydrogels formed by specific protein-peptide interactions. [6] Specific protein-peptide interaction has also been used to direct selfassembly of peptide nanowires into micrometer-sized crystalline cubes. [7] However, there are no reports about the formation of molecular hydrogels through protein-peptide interactions up to now. As mentioned above, the formation of a 3D matrix is crucial to the formation of molecular hydrogels. To use protein-peptide interactions to enhance interfiber interactions to support 3D structures, fusion proteins with multiple binding sites are needed. These kinds of fusion proteins usually contain two parts, one for multimer formation and the other for peptide binding. However, these fusion proteins are usually in the balance between multimers with multiple binding sites and monomer with only one binding site. The dissociation constants of protein-peptide interactions are also usually in the micro-to millimolar range. It remains a challenge to develop a protein that can predominantly (> 95 %) form multimers with multiple binding sites. It would also be highly interesting for researchers in the field of biomaterials to develop a protein-peptide interaction with Figure 1. Protein-peptide interaction can be used to enhance interactions between self-assembled fibers, thus leading to molecular hydrogelation. A) Chemical structure of Nap-GFFYGGGWRESAI (1: nongelator) with a possible self-assembly a...

Self-Assembly-Induced Far-Red/Near-Infrared Fluorescence Light-Up for Detecting and Visualizing Specific Protein–Peptide Interactions

“…TIP-1 and ULD-TIP-1 proteins were expressed and purified using standard recombinant protein technology as previously reported. 38,39 In brief, DNA fragments corresponding to protein sequences were cloned into an in-house modified version of the pET32a (Novagen) in which the S-tag and the thrombin recognition site were replaced with a sequence encoding a PreScission protease-cleavable segment (Leu-Glu-Val-Leu-Phe-Gln-Gly-Pro). The resulting protein contained a Trx-His 6 -tag in its N-terminus.…”

“…38 Moreover, ULD protein can spontaneously form a tetramer, making ULD-TIP-1 appear as a polyprotein with four binding sites to peptides. 39 As protein aggregation is associated with several diseases such as Alzheimer's and Prion diseases, 40,41 detection of polyprotein and polyproteinÀ peptide interactions is of vital importance. In this work, the ULD-TIP-1 tetramer was used as a model polyprotein to assess the feasibility of our probe for detection of polyproteinÀpeptide interactions.…”

Supramolecular Guest–Host Interactions for the Preparation of Biomedical Materials

“…These include the relatively weak association of crown ethers, cryptands, and related molecules; [23][24] though, such associations are generally more suited for ionic sequestration and not molecular recognition. Alternatively, an array of transient peptide and/or protein interactions have been devised, [25][26][27][28][29] which often display greater affinity and specificity, but at the cost of increasing complexity and laborious synthesis. Finally, the guest-host macrocycles combine rapid, high affinity molecular recognition with ease and scalability of synthesis.…”

Mixing Biomimetic Heterodimers of Nucleopeptides to Generate Biocompatible and Biostable Supramolecular Hydrogels

“…This approach, in fact, has been explored by a few groups by using peptides to bind with proteins. [15] For example, the specific TPR-peptide interaction, [15a] TIP1-peptide interaction, [15b] heparin-VEGF interaction, [15c] allows the formation of polymeric hydrogels. One drawback of this approach is the use of proteins being high cost and susceptible to proteolysis.…”