Protein aggregation – quantifying platform
Protein aggregation is a hallmark of aging and amyloid diseases. At the same time, it constitutes an important issue in the manufacturing process of biopharmaceuticals. Being able to promptly detect soluble aggregates before mature fibrils form would help intervene early in the manufacture process and avoid potential toxicity issues. The EU-funded PROARGUS (Protein aggregation - a quantitative assessment) project set out to develop a sensing platform dedicated to detecting soluble aggregates. They exploited molecular imprinted polymers to generate sensors that can detect protein monomers, dimers and more complex aggregates. The consortium used a number of analytical methods including size exclusion chromatography and matrix assisted laser desorption ionisation time of flight mass spectrometry to monitor the aggregation process. The progressive stages from unfolding and oligomerisation to the formation of mature fibrils were screened using lysozyme as a model system. Next, researchers developed a platform for quantitative assessment of aggregation. They incorporated a novel aptamer and surface plasmon resonance-based sensor that could detect the onset of aggregation by distinguishing between lysozyme oligomers and monomers. The sensor was tested in real-life situations such as wine where the presence of lysozymes can trigger allergic reactions. Following optimisation, it was possible to detect, 0.1 % lysozyme dimer in a monomer solution. Overall, the PROARGUS platform proved to be a sensitive tool for the detection of protein oligomers. Although it was developed and tested on lysozymes, its utilisation can be extended in the future to therapeutic proteins/peptides, for which specific recognition elements exist.
Keywords
Protein aggregation, manufacture process, sensor, lysozyme
