The primary structure of a protein consists of a sequence of amino acids and is a key factor in determining how a protein folds and functions. However, conventional methods for sequencing proteins, such as mass spectrometry and Edman degradation, suffer from short reads and lack sensitivity, so alternative approaches are sought. Here, we show that a subnanometre-diameter pore, sputtered through a thin silicon nitride membrane, can be used to detect the primary structure of a denatured protein molecule. When a denatured protein immersed in electrolyte is driven through the pore by an electric field, measurements of a blockade in the current reveal nearly regular fluctuations, the number of which coincides with the number of residues in the protein. Furthermore, the amplitudes of the fluctuations are highly correlated with the volumes that are occluded by quadromers (four residues) in the primary structure. Each fluctuation, therefore, represents a read of a quadromer. Scrutiny of the fluctuations reveals that the subnanometre pore is sensitive enough to read the occluded volume that is related to post-translational modifications of a single residue, measuring volume differences of ∼0.07 nm, but it is not sensitive enough to discriminate between the volumes of all twenty amino acids.
Worldwide, as threats from chemical warfare (CW) agents have increased, the need for sensors and detectors for such agents has also risen. Many of the most dangerous CW agents are covalent inhibitors of acetylcholinesterase (AchE) which function by very similar chemistry, and hence have similar chemical properties overall. Here we show that a recently developed class of fluorescent molecules with phenylene ethynylene (PE) backbones can act as excellent sensors for a class of CW simulant molecules (malathion and its derivatives) when in complex with an anionic surfactant at very low concentrations. Oligo phenylene ethynylenes (OPEs) have recently been shown to be both efficient fluorophores and (depending on details of structure) sources of photo-generated reactive oxygen species, especially singlet oxygen. As such, the OPEs are attractive candidates as both potential CW sensors and potential CWdestroying agents. Here we show that several OPE variants are able to detect the CW simulant malathion when in aqueous complex with small concentrations of surfactants such as sodium dodecyl sulfate (SDS). At the correct molar ratios, the combination of the OPE molecule with SDS and malathion appears to form a cooperative triple complex that varies sharply in its fluorescence properties over a narrow range of malathion concentrations. With some OPE/ surfactant combinations the complex acts as a ''turn off'' detector-fluorescence intensity decreases with malathion concentration-while other combinations can act as ''turn on'' detectors-fluorescence intensity increases with malathion concentration. Simulations designed to understand the nature of the triple complex, and further experiments to test the ability of triple complexes to destroy CW agents, are currently underway.
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