Zeyuan Dong

Construction of GPx Active Centers on Natural Protein Nanodisk/Nanotube: A New Way to Develop Artificial Nanoenzyme

Construction of catalytic centers on natural protein aggregates is a challenging topic in biomaterial and biomedicine research. Here we report a novel construction of artificial nanoenzyme with glutathione peroxidase (GPx)-like function. By engineering the surface of tobacco mosaic virus (TMV) coat protein, the main catalytic components of GPx were fabricated on TMV protein monomers. Through direct self-assembly of the functionalized viral coat proteins, the multi-GPx centers were installed on these well-defined nanodisks or nanotubes. With the help of muti-selenoenzyme centers, the resulting organized nanoenzyme exhibited remarkable GPx activity, even approaching the level of natural GPx. The antioxidation study on subcell mitochondrial level demonstrated that virus-based nanoenzyme exerted excellent capacity for protecting cell from oxidative damage. This strategy represents a new way to develop artificial nanoenzymes.

Self-assembly of amphiphilic peptides into bio-functionalized nanotubes: a novel hydrolase model

Herein, we report the construction of a novel hydrolase model via self-assembly of a synthetic amphiphilic short peptide (Fmoc-FFH-CONH2) into nanotubes. The peptide-based self-assembled nanotubes (PepNTs-His) with imidazolyl groups as the catalytic centers exhibit high catalytic activity for p-nitrophenyl acetate (PNPA) hydrolysis. By replacement of the histidine of Fmoc-FFH-CONH2 with arginine to produce a structurally similar peptide Fmoc-FFR-CONH2, guanidyl groups can be presented in the nanotubes through the co-assembly of these two molecules to stabilize the transition state of the hydrolytic reaction. Therefore significantly improved catalytic activity has been achieved by the reasonable distribution of three dominating catalytic factors: catalytic center, binding site and transition state stabilization to the co-assembled peptide nanotubes (PepNTs-His-Argmax). The resulting hydrolase model shows typical saturation kinetics behaviour to that of natural enzymes and the catalytic efficiency of a single catalytic center is 519-fold higher than that without catalysts. As for a nanotube with multi-catalytic centers, a remarkable catalytic efficiency could be achieved with the increase of building blocks. This model suggests that the well ordered and dynamic supramolecular structure is an attractive platform to develop new artificial enzymes to enhance the catalytic activity. Besides, this novel peptide-based material has excellent biocompatibility with human cells and is expected to be applied to organisms as a substitute for natural hydrolases.

Enzymetically Regulating the Self‐Healing of Protein Hydrogels with High Healing Efficiency

Enzyme-mediated self-healing of dynamic covalent bond-driven protein hydrogels was realized by the synergy of two enzymes, glucose oxidase (GOX) and catalase (CAT). The reversible covalent attachment of glutaraldehyde to lysine residues of GOX, CAT, and bovine serum albumin (BSA) led to the formation and functionalization of the self-healing protein hydrogel system. The enzyme-mediated protein hydrogels exhibit excellent self-healing properties with 100% recovery. The self-healing process was reversible and effective with an external glucose stimulus at room temperature.

Dual stimuli-responsive supramolecular pseudo-polyrotaxane hydrogels

THPP-(PEG2000-BA)4, a four-branched molecule end-decorated with benzaldehyde (BA), was successfully designed and synthesized. It can form physical pseudo-polyrotaxane (PPR) hydrogels in the presence of α-cyclodextrins (α-CDs). The branched structure of the THPP core promotes gel formation with a small amount of host and guest. Moreover, these resulting hydrogels are dual stimuli-responsive, which can be observed by physically macroscopical phenomena and 1H NMR spectra. Since BA can react with amine to form a pH-responsive Schiff-base which possesses a dynamic covalent nature, it is anticipated that the formation of gels can be modulated by pH. When THPP-(PEG2000-BA)4 was blocked by the relatively large molecule 6-N-ethylenediamine-6-deoxy functionalized β-CD (EDA-β-CD) gelation did not take place. However, the addition of an acid resulted in gelation since the benzoic imine bonds can hydrolyze under acidic conditions and α-CDs can thus pass through BA to thread on the polyethylene glycol (PEG) chains. When a strongly competitive guest, 1-[p-(phenylazo)benzyl]-bromide (Azo-C1-N+), was added to the gel, a gel-to-sol transition was observed due to the disassembly of inclusion complexes between α-CDs and PEG chains. Simultaneously, this gel shows a photo-responsive capacity because of the presence of azobenzene. Therefore, the cycles of gel–sol transitions were achieved through pH- and photo-stimuli. This kind of hydrogel is promising for use in many fields, such as biology and electronics.

Temperature-driven switching of the catalytic activity of artificial glutathione peroxidase by the shape transition between the nanotubes and vesicle-like structures.

Smart supramolecular nanoenzymes with temperature-driven switching property have been successfully constructed by the self-assembly of supra-amphiphiles formed by the cyclodextrin-based host-guest chemistry. The self-assembled nanostructures were catalyst-functionalized and thermosensitively-functionalized through conveniently linking the catalytic center of glutathione peroxidase and thermosensitive polymer to the host cyclodextrin molecules.The ON-OFF switches for the peroxidase activity by reversible transformation of nanostructures from tube to sphere have been achieved through changing the temperature. We anticipate that such intelligent enzyme mimics could be developed to use in an antioxidant medicine with controlled catalytic efficiency according to the needs of the human body in the future.

Reversible Ca2+ Switch of An Engineered Allosteric Antioxidant Selenoenzyme†

A Ca(2+) -responsive artificial selenoenzyme was constructed by computational design and engineering of recoverin with the active center of glutathione peroxidase (GPx). By combining the recognition capacity for the glutathione (GSH) substrate and the steric orientation of the catalytic selenium moiety, the engineered selenium-containing recoverin exhibits high GPx activity for the catalyzed reduction of H2 O2 by glutathione (GSH). Moreover, the engineered selenoenzyme can be switched on/off by Ca(2+) -induced allosterism of the protein recoverin. This artificial selenoenzyme also displays excellent antioxidant ability when it was evaluated using a mitochondrial oxidative damage model, showing great potential for controlled catalysis in biomedical applications.

Biomimetic Transmembrane Channels with High Stability and Transporting Efficiency from Helically Folded Macromolecules.

Membrane channels span the cellular lipid bilayers to transport ions and molecules into cells with sophisticated properties including high efficiency and selectivity. It is of particular biological importance in developing biomimetic transmembrane channels with unique functions by means of chemically synthetic strategies. An artificial unimolecular transmembrane channel using pore-containing helical macromolecules is reported. The self-folding, shape-persistent, pore-containing helical macromolecules are able to span the lipid bilayer, and thus result in extraordinary channel stability and high transporting efficiency for protons and cations. The lifetime of this artificial unimolecular channel in the lipid bilayer membrane is impressively long, rivaling those of natural protein channels. Natural channel mimics designed by helically folded polymeric scaffolds will display robust and versatile transport-related properties at single-molecule level.

A Dual Enzyme Microgel with High Antioxidant Ability Based on Engineered Seleno‐Ferritin and Artificial Superoxide Dismutase

An antioxidant microgel with both glutathione peroxidase (GPx) and superoxide dismutase (SOD) activities is reported. Using computational design and genetic engineering methods, the main catalytic components of GPx are fabricated onto the surface of ferritin. The resulting seleno-ferritin (Se-Fn) monomers can self-assemble into nanocomposites that exhibit remarkable GPx activity due to the well organized multi-GPx catalytic centers. Subsequently, a porphyrin derivative is synthesized as an SOD mimic, and is employed to construct a synergistic dual enzyme system by crosslinking Se-Fn nanocomposites into a microgel. Significantly, this dual enzyme microgel is demonstrated to display better antioxidant ability than single GPx or SOD mimics in protecting cells from oxidative damage.

Design of aromatic helical polymers for STM visualization: imaging of single and double helices with a pattern of π-π stacking.

From scanning tunneling microscopy (STM) images of rationally designed helical polymers with a pattern of π-π stacking, we successfully identified the single- and double-helical superstructures. The STM images of the helical structures revealed the smallest helical architecture (diameter ca. 1.3 nm) that has been seen so far. Furthermore, the interconversion of single and double helices was further underpinned by experimental analyses. Significantly, the formation of double helices induced different supramolecular chirality to that observed for the single helices.

Understanding enzyme catalysis by means of supramolecular artificial enzymes

Enzymes are biomacromolecules responsible for the abundant chemical biotransformations that sustain life. Recently, biochemists have discovered that multiple conformations and numerous parallel paths are involved during the processes catalyzed by enzymes. It is plausible that the entire macromolecular scaffold is involved in catalysis via cooperative motions that result in incredible catalytic efficiency. Moreover, some enzymes can very strongly bind the transition state with an association constant of up to 1024 M−1, suggesting that covalent bond formation is a possible process during the conversion of the transition state in enzyme catalysis, in addition to the concatenation of non-covalent interactions. Supramolecular chemistry provides fundamental knowledge about the relationships between the dynamic structures and functions of organized molecules. By taking advantage of supramolecular concepts, numerous supramolecular enzyme mimics with complex and hierarchical structures have been designed and investigated. Through the study of supramolecular enzyme models, a great deal of information to aid our understanding of the mechanism of catalysis by natural enzymes has been acquired. With the development of supramolecular artificial enzymes, it is possible to replicate the features of natural enzymes with regards to their constitutional complexity and cooperative motions, and eventually decipher the conformation-based catalytic mystery of natural enzymes.