A plug-and-play aptamer diagnostic platform based on linear dichroism spectroscopy.

A plug-and-play sandwich assay platform for the aptamer-based detection of molecular targets using linear dichroism (LD) spectroscopy as a read-out method has been demonstrated. A 21-mer DNA strand comprising the plug-and-play linker was bioconjugated onto the backbone of the filamentous bacteriophage M13, which gives a strong LD signal due to its ready alignment in linear flow. Extended DNA strands containing aptamer sequences that bind the protein thrombin, TBA and HD22, were then bound to the plug-and-play linker strand via complementary base pairing to generate aptamer-functionalised M13 bacteriophages.

Combining bacteriophage engineering and linear dichroism spectroscopy to produce a DNA hybridisation assay.

Nucleic acid detection is an important part of our bio-detection arsenal, with the COVID-19 pandemic clearly demonstrating the importance to healthcare of rapid and efficient detection of specific pathogenic sequences. As part of the drive to establish new DNA detection methodologies and signal read-outs, here we show how linear dichroism (LD) spectroscopy can be used to produce a rapid and modular detection system for detecting quantities of DNA from both bacterial and viral pathogens. The LD sensing method exploits changes in fluid alignment of bionanoparticles (bacteriophage M13) engineered with DNA stands covalently attached to their surfaces, with the read-out signal induced by the formation of complementary duplexes between DNA targets and two M13 bionanoparticles.

Ligand-induced conformational changes in a SMALP-encapsulated GPCR.

The adenosine 2A receptor (AR), a G-protein-coupled receptor (GPCR), was solubilised and purified encapsulated in styrene maleic acid lipid particles (SMALPs). The purified AR-SMALP was associated with phospholipids characteristic of the plasma membrane of Pichia pastoris, the host used for its expression, confirming that the AR-SMALP encapsulated native lipids. The fluorescence spectrum of the AR-SMALP showed a characteristic broad emission peak at 330 nm, produced by endogenous Trp residues.

Light-controlled thrombin catalysis and clot formation using a photoswitchable G-quadruplex DNA aptamer.

The reversible photocontrol of an enzyme governing blood coagulation is demonstrated. The thrombin binding aptamer (TBA), was rendered photochromic by modification with two anthracene groups. Light-triggered anthracene photodimerisation distorts its structure, inhibiting binding of the enzyme thrombin, which in turn triggers catalysis and the resulting clotting process.

Linear dichroism of visible-region chromophores using M13 bacteriophage as an alignment scaffold.

It is a challenge within the field of biomimetics to recreate the properties of light-harvesting antennae found in plants and photosynthetic bacteria. Attempts to recreate these biological structures typically rely on the alignment of fluorescent moieties attachment to an inert linear scaffold, DNA, RNA or amyloid fibrils, to enable Förster resonance energy transfer (FRET) between attached chromophores. While there has been some success in this approach, refinement of the alignment of the chromophores is often limited, which may limit the efficiency of energy transfer achieved.

Dye Aggregate-Mediated Self-Assembly of Bacteriophage Bioconjugates.

One of the central themes of biomolecular engineering is the challenge of exploiting the properties of biological materials. Part of this challenge has been uncovering and harnessing properties of biological components that only emerge following their ordered self-assembly. One biomolecular building block that has received significant interest in the past decade is the M13 bacteriophage.

Direct detection and measurement of wall shear stress using a filamentous bio-nanoparticle.

The wall shear stress (WSS) that a moving fluid exerts on a surface affects many processes including those relating to vascular function. WSS plays an important role in normal physiology (e.g.

Polymerase Chain Reaction on a Viral Nanoparticle.

The field of synthetic biology includes studies that aim to develop new materials and devices from biomolecules. In recent years, much work has been carried out using a range of biomolecular chassis including α-helical coiled coils, β-sheet amyloids and even viral particles. In this work, we show how hybrid bionanoparticles can be produced from a viral M13 bacteriophage scaffold through conjugation with DNA primers that can template a polymerase chain reaction (PCR).