Experimental and numerical evaluation of a genetically engineered M13 bacteriophage with high sensitivity and selectivity for 2,4,6-trinitrotoluene.

Selective and sensitive detection of desired targets is very critical in sensor design. Here, we report a genetically engineered M13 bacteriophage-based sensor system evaluated by quantum mechanics (QM) calculations. Phage display is a facile way to develop the desired peptide sequences, but the resulting sequences can be imperfect peptides for binding of target molecules. A TNT binding peptide (WHW) carrying phage was self-assembled to fabricate thin films and tested for the sensitive and selective surface plasmon resonance-based detection of TNT molecules at the 500 femtomole level. SPR studies performed with the WHW peptide and control peptides (WAW, WHA, AHW) were well-matched with those of the QM calculations. Our combined method between phage engineering and QM calculation will significantly enhance our ability to design selective and sensitive sensors.

Selective and Sensitive Sensing of Flame Retardant Chemicals Through Phage Display Discovered Recognition Peptide.

We report a highly selective and sensitive biosensor for the detection of an environmentally toxic molecule, decabrominated diphenyl ether (DBDE), one of the most common congeners of the polybrominated frame retardants (polybrominated diphenyl ether (PBDE)), using newly discovered DBDE peptide receptors integrated with carbon nanotube field-effect transistors (CNT-FET). The specific DBDE peptide receptor was identified using a high-throughput screening process of phage library display. The resulting binding peptide carries an interesting consensus binding pocket with two Trp-His/Asn-Trp repeats, which binds to the DBDE in a multivalent manner. We integrated the novel DBDE binding peptide onto the CNT-FET using polydiacetylene coating materials linked through cysteine-maleimide click chemistry. The resulting biosensor could detect the desired DBDE selectively with a 1 fM detection limit. Our combined approaches of selective receptor discovery, material nanocoating through click chemistry, and integration onto a sensitive CNT-FET electronic sensor for desired target chemicals will pave the way toward the rapid development of portable and easy-to-use biosensors for desired chemicals to protect our health and environment.

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.

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.

Biomimetic virus-based colourimetric 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.

Virus-based piezoelectric energy generation.

Piezoelectric materials can convert mechanical energy into electrical energy, and piezoelectric devices made of a variety of inorganic materials and organic polymers have been demonstrated. However, synthesizing such materials often requires toxic starting compounds, harsh conditions and/or complex procedures. Previously, it was shown that hierarchically organized natural materials such as bones, collagen fibrils and peptide nanotubes can display piezoelectric properties. Here, we demonstrate that the piezoelectric and liquid-crystalline properties of M13 bacteriophage (phage) can be used to generate electrical energy. Using piezoresponse force microscopy, we characterize the structure-dependent piezoelectric properties of the phage at the molecular level. We then show that self-assembled thin films of phage can exhibit piezoelectric strengths of up to 7.8 pm V(-1). We also demonstrate that it is possible to modulate the dipole strength of the phage, hence tuning the piezoelectric response, by genetically engineering the major coat proteins of the phage. Finally, we develop a phage-based piezoelectric generator that produces up to 6 nA of current and 400 mV of potential and use it to operate a liquid-crystal display. Because biotechnology techniques enable large-scale production of genetically modified phages, phage-based piezoelectric materials potentially offer a simple and environmentally friendly approach to piezoelectric energy generation.