Seung-Wuk Lee

Biomimetic self-templating supramolecular structures

In nature, helical macromolecules such as collagen, chitin and cellulose are critical to the morphogenesis and functionality of various hierarchically structured materials. During tissue formation, these chiral macromolecules are secreted and undergo self-templating assembly, a process whereby multiple kinetic factors influence the assembly of the incoming building blocks to produce non-equilibrium structures. A single macromolecule can form diverse functional structures when self-templated under different conditions. Collagen type I, for instance, forms transparent corneal tissues from orthogonally aligned nematic fibres, distinctively coloured skin tissues from cholesteric phase fibre bundles, and mineralized tissues from hierarchically organized fibres. Nature’s self-templated materials surpass the functional and structural complexity achievable by current top-down and bottom-up fabrication methods. However, self-templating has not been thoroughly explored for engineering synthetic materials. Here we demonstrate the biomimetic, self-templating assembly of chiral colloidal particles (M13 phage) into functional materials. A single-step process produces long-range-ordered, supramolecular films showing multiple levels of hierarchical organization and helical twist. Three distinct supramolecular structures are created by this approach: nematic orthogonal twists, cholesteric helical ribbons and smectic helicolidal nanofilaments. Both chiral liquid crystalline phase transitions and competing interfacial forces at the interface are found to be critical factors in determining the morphology of the templated structures during assembly. The resulting materials show distinctive optical and photonic properties, functioning as chiral reflector/filters and structural colour matrices. In addition, M13 phages with genetically incorporated bioactive peptide ligands direct both soft and hard tissue growth in a hierarchically organized manner. Our assembly approach provides insight into the complexities of hierarchical assembly in nature and could be expanded to other chiral molecules to engineer sophisticated functional helical-twisted structures.

Light-Controlled Graphene-Elastin Composite Hydrogel Actuators

Hydrogels actuators (HAs) that can reversibly respond to stimuli have applications in diverse fields. However, faster response rates and improved control over actuation timing and location are required to fulfill their potential. To address these criteria, we synthesized near-infrared light-driven HAs by interfacing genetically engineered elastin-like polypeptides with reduced-graphene oxide sheets. The resulting nanocomposites exhibited rapid and tunable motions controlled by light position, intensity, and path, including finger-like flexing and crawling. This work demonstrates the ability of rationally designed proteins to be combined with synthetic nanoparticles for the creation of macroscale functional materials.

Genetically engineered nanofiber-like viruses for tissue regenerating materials.

Controlling structural organization and signaling motif display of biomimetic matrices at the nanometer scale is of great importance to the functional design of tissue regenerating materials. We have genetically engineered M13 bacteriophage (phage), naturally occurring nanofiber-like viruses, to display a high density of cell-signaling peptides on their major coat proteins. Structural orientation of these phage building blocks can be achieved due to their long-rod shape and monodispersity, which lead them to self-assemble into directionally organized liquid crystalline-like materials. We showed that the constructed viral nanofiber scaffolds were able to support neural progenitor cell proliferation and differentiation as well as direct orientation of their growth in three dimensions. Such functionalized and structurally aligned phage matrices offer promising opportunities for therapies that address challenging medical problems, such as nerve tissue regeneration after spinal cord injuries, or as in vitro model systems for studying complicated cell signaling environments.

Evolutionary Screening of Biomimetic Coatings for Selective Detection of Explosives

Susceptibility of chemical sensors to false positive signals remains a common drawback due to insufficient sensor coating selectivity. By mimicking biology, we have demonstrated the use of sequence-specific biopolymers to generate highly selective receptors for trinitrotoluene and 2,4-dinitrotoluene. Using mutational analysis, we show that the identified binding peptides recognize the target substrate through multivalent binding with key side chain amino acid elements. Additionally, our peptide-based receptors embedded in a hydrogel show selective binding to target molecules in the gas phase. These experiments demonstrate the technique of receptor screening in liquid to be translated to selective gas-phase target binding, potentially impacting the design of a new class of sensor coatings.

Biomimetic virus-based colorimetric sensors

Many materials in nature change colours in response to stimuli, making them attractive for use as sensor platform. However, both natural materials and their synthetic analogues lack selectivity towards specific chemicals, and introducing such selectivity remains a challenge. Here we report the self-assembly of genetically engineered viruses (M13 phage) into target-specific, colourimetric biosensors. The sensors are composed of phage-bundle nanostructures and exhibit viewing-angle independent colour, similar to collagen structures in turkey skin. On exposure to various volatile organic chemicals, the structures rapidly swell and undergo distinct colour changes. Furthermore, sensors composed of phage displaying trinitrotoluene (TNT)-binding peptide motifs identified from a phage display selectively distinguish TNT down to 300 p.p.b. over similarly structured chemicals. Our tunable, colourimetric sensors can be useful for the detection of a variety of harmful toxicants and pathogens to protect human health and national security.

Selective and Sensitive TNT Sensors Using Biomimetic Polydiacetylene- Coated CNT-FETs

Miniaturized smart sensors that can perform sensitive and selective real-time monitoring of target analytes are tremendously valuable for various sensing applications. We developed selective nanocoatings by combining trinitrotoluene (TNT) receptors bound to conjugated polydiacetylene (PDA) polymers with single-walled carbon nanotube field-effect transistors (SWNT-FET). Selective binding events between the TNT molecules and phage display derived TNT receptors were effectively transduced to sensitive SWNT-FET conductance sensors through the PDA coating layers. The resulting sensors exhibited an unprecedented 1 fM sensitivity toward TNT in real time, with excellent selectivity over various similar aromatic compounds. Our biomimetic receptor coating approach may be useful for the development of sensitive and selective micro- and nanoelectronic sensor devices for various other target analytes.

Polymer-oligopeptide composite coating for selective detection of explosives in water.

The selective detection of a specific target molecule in a complex environment containing potential contaminants presents a significant challenge in chemical sensor development. Utilizing phage display techniques against trinitrotoluene (TNT) and dinitrotoluene (DNT) targets, peptide receptors have previously been identified with selective binding capabilities for these molecules. For practical applications, these receptors must be immobilized onto the surface of sensor platforms at high density while maintaining their ability to bind target molecules. In this paper, a polymeric matrix composed of poly(ethylene-co-glycidyl methacrylate) (PEGM) has been prepared. A high density of receptors was covalently linked through reaction of amino groups present in the receptor with epoxy groups present in the co-polymer. Using X-ray photoelectron spectroscopy (XPS) and gas-chromatography/mass spectroscopy (GC/MS), this attachment strategy is demonstrated to lead to stably bound receptors, which maintain their selective binding ability for TNT. The TNT receptor/PEGM conjugates retained 10-fold higher TNT binding ability in liquid compared to the lone PEGM surface and 3-fold higher TNT binding compared to non-specific receptor conjugates. In contrast, non-target DNT exposure yielded undetectable levels of binding. These results indicate that this polymeric construct is an effective means of facilitating selective target interaction both in an aqueous environment. Finally, real-time detection experiments were performed using a quartz crystal microbalance (QCM) as the sensing platform. Selective detection of TNT vs DNT was demonstrated using QCM crystals coated with PEGM/TNT receptor, highlighting that this receptor coating can be incorporated as a sensing element in a standard detection device for practical applications.

Polydiacetylene incorporated with peptide receptors for the detection of trinitrotoluene explosives.

Because of their unique optical and stimuli-response properties, polydiacetylene-based platforms have been explored as an alternative to complex mechanical and electrical sensing systems. We linked chromic responsive polydiacetylene (PDA) onto a peptide-based molecular recognition element for trinitrotoluene (TNT) molecules in order to provide a system capable of responding to the presence of a TNT target. We first identified the trimer peptide receptor that could induce chromic changes on a PDA backbone. We then investigated the multivalent interactions between TNT and our peptide-based receptor by nuclear magnetic resonance (NMR) spectroscopy. We further characterized various parameters that affected the conjugated PDA system and hence the chromic response, including the size of end-group motifs, the surface density of receptors, and the length of alkane side chains. Taking these necessary design parameters into account, we demonstrated a modular system capable of transducing small-molecule TNT binding into a detectable signal. Our conjugated PDA-based sensor coupled with molecular recognition elements has already proven useful recently in the development of another sensitive and selective electronic sensor, though we expect that our results will also be valuable in the design of colorimetric sensors for small-molecule detection.

Assembly of Bacteriophage into Functional Materials

For the last decade, the fabrication of ordered structures of phage has been of great interest as a means of utilizing the outstanding biochemical properties of phage in developing useful materials. Combined with other organic/inorganic substances, it has been demonstrated that phage is a superior building block for fabricating various functional devices, such as the electrode in lithium-ion batteries, photovoltaic cells, sensors, and cell-culture supports. Although previous research has expanded the utility of phage when combined with genetic engineering, most improvements in device functionality have relied upon increases in efficiency owing to the compact, more densely packable unit size of phage rather than on the unique properties of the ordered nanostructures themselves. Recently, self-templating methods, which control both thermodynamic and kinetic factors during the deposition process, have opened up new routes to exploiting the ordered structural properties of hierarchically organized phage architectures. In addition, ordered phage films have exhibited unexpected functional properties, such as structural color and optical filtering. Structural colors or optical filtering from phage films can be used for optical phage-based sensors, which combine the structural properties of phage with target-specific binding motifs on the phage-coat proteins. This self-templating method may contribute not only to practical applications, but also provide insight into the fundamental study of biomacromolecule assembly in in vivo systems under complicated and dynamic conditions.

Evolutionary screening of collagen-like peptides that nucleate hydroxyapatite crystals.

The biogenesis of inorganic/organic composite materials such as bone typically involves the process of templated mineralization. Biomimetic synthesis of bone-like materials therefore requires the development of organic scaffolds that mediate mineralization of hydroxyapatite (HAP), the major inorganic component of bone. Using phage display, we identified a 12-residue peptide that bound to single-crystal HAP and templated the nucleation and growth of crystalline HAP mineral in a sequence- and composition-dependent manner. The sequence responsible for the mineralizing activity resembled the tripeptide repeat (Gly-Pro-Hyp) of type I collagen, a major component of bone extracellular matrix. Using a panel of synthetic peptides, we defined the structural features required for mineralizing activity. The results support a model for the cooperative noncovalent interaction of the peptide with HAP and suggest that native collagen may have a mineral-templating function in vivo. We expect this short HAP-binding peptide to be useful in the synthesis of three-dimensional bone-like materials.