Single molecule protein sequencing enables identification of the primary structure of biologically active peptides and proteins. This is relevant for their application as food products, but also to better understand and fight diseases. Already small amounts of modified protein can lead to unwanted flavors in food proteins, but also to serious diseases such as cancer. Although the new nanopore sequencing technology allows determination of a sequence of small amounts of protein, it cannot process intact peptides and proteins. We will develop a robust bench-compatible method to convert a biological peptide or protein into DNA-peptide-DNA sandwich conjugates that can be analyzed by nanopores on a single molecule level. For this, we use established enzymes to digest a variety of biological peptides or proteins (and their mixtures), and follow this up with robust and orthogonal ligation techniques. Our process does not rely on expensive analytical equipment (pipettes and a centrifuge should suffice), and should only use water and aqueous buffers, thereby limiting the amount of chemical reagents. As such, our bench-compatible method avoids the involvement of a dedicated and exclusive specialized lab. This enables widespread application, also in less wealthy countries, in areas such as food production.
