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Development of protein nanopores for their application in biotechnology and medicine

Nanopore technology has advanced rapidly over the past three decades, demonstrating its potential for detecting various biomolecules. This technology uses a nanometer-sized aperture (a nanopore) embedded in a biological or artificial support that connects two reservoirs filled with an electrolyte solution. When a voltage is applied, ions move through the nanopore and create current. Due to electrophoretic or electroosmotic forces, individual analyte molecules are forced to pass through the pore, and the partial blockage of the pore is registered as a change in electric current. Each molecule can be characterized by its current signature (i.e. the specific change in the current and the time it takes to pass from one chamber to the other), which allows us to monitor the presence of different molecules at the single-molecule level.

In our department, we are interested in pore-forming toxins (PFTs), which are well suited for nanopore applications. These naturally occurring proteins form nanometer-sized pores through a lipid membrane. The pore size is well defined, but can range from 1-2 nm to 40 nm depending on the toxin family. In addition to the defined pore size, the unique architecture and shape of the transmembrane region are further arguments why natural pores are ideal candidates for nanopore applications. Our efforts are aimed at identifying novel PFTs from recently deposited genomic and metagenomic sources, characterizing recombinantly expressed nanopores, and fine-tuning their properties through directed evolution.

The wide range of PFT sizes allows us to adapt them for the detection of analytes of different sizes. We are particularly interested in the detection of various proteins, focusing on human proteins that have high prognostic or diagnostic value. Because nanopore technology enables real-time, label-free detection from tiny sample volumes, nanopore-based detection is becoming an attractive alternative for protein detection in place of traditional methods such as mass spectrometry.

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