Development of Protoporphyrin Derivatives as Potential Amyloid Inhibitors

Gayen, Soumajit (2022) Development of Protoporphyrin Derivatives as Potential Amyloid Inhibitors. Masters thesis, Indian Institute of Science Education and Research Kolkata.

Text (MS dissertation of Soumajit Gayen (17MS208)) 17MS208_Thesis_file.pdf - Submitted Version Restricted to Repository staff only Download (3MB)

Abstract

Protein misfolding and toxic aggregation, commonly referred to as amyloids, and their deposition in functionally important regions of our body has been associated with many chronic and fatal neurodegenerative diseases (amyloidosis). A protein of our own body, due to certain triggering conditions, gets transformed from its native state to β-sheet rich fibres and finally to insoluble toxic aggregates – the plaques, the root cause of these diseases. The ‘impossible to cure’ diseases like Alzheimer’s, Parkinson’s etc. owe their existence to these unwanted depositions, moreover, proteins like IAPP and insulin and its amyloidosis have been linked to some common chronic diseases like diabetes. Once the fibres are formed and deposited inside the body it is near impossible to degrade it. Thus, finding a solution to the undesirable aggregation is an urgency. In order to aid the cause and to find a therapeutically relevant molecule in inhibiting these toxic protein aggregations, we have synthesized a library of Protoporphyrin derivatives (PPDs) and studied its efficiency in inhibition of in-vitro fibrillization of insulin and lysozyme. Upon monitoring the fibrillization kinetics in presence of PPDs, it was noted that fibrillization of both insulin and lysozyme was significantly reduced and even completely arrested. It was duly noted that with the introduction of groups capable of modulating or enhancing non-covalent interactions, the inhibition efficiency increased. Although the relationship is not exactly linear, it was inferred that the efficacy of these compounds and the way they interact with the protein depends on the protein’s structure as well. Evidence from the primary ThT based fluorescence suggested that i-BHP (the O-isobutyl derivative of Hematoporphyrin) completely arrested the fibrillization of insulin upto 100 hours, which was confirmed by scanning electron microscopy (FESEM) and circular dichroism spectroscopy (CD). The results from the finer experiments suggested that the conformational change of the proteins (from α-helix to β-sheet) was probably halted in presence of the PPDs, CD analysis confirmed the preservation of the native α-helix structure in samples incubated with PPDs after the end of the experimentation time-frame. Thus, the synthesized molecules have the potential to be implemented as effective amyloid inhibitors after due tests and screenings and experimentation with other relevant proteins. The promising observations calls for an understanding on the details of the intricate mechanism of protein misfolding and how our molecules show its effect, which shall be undertaken in near future. Finally, these results may provide a useful insight in developing new possible therapeutical approaches towards the prevention of amyloidosis.

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