Employing in vitro analysis to test the potency of methylglyoxal in inducing the formation of amyloid-like aggregates of caprine brain cystatin

Thiol protease inhibitors (cystatins) are implicated in various disease states from cancer to neurodegenerative conditions and immune responses. Cystatins have high amyloidogenic propensity and they are prone to form fibrillar aggregates leading to amyloidosis. Particularly challenging examples of such disorders occur in type 2 diabetes, Alzheimer's and Parkinson's diseases. The aim of the present study is to find an interaction between the compound methylglyoxal (MG) which is particularly elevated in type 2 diabetes with caprine brain cystatin (CBC). Results have shown that elevated concentration of MG forms amyloid aggregates of CBC. This was achieved by allowing slow growth in a solution containing moderate to high concentrations of MG. When analysed with microscopy, the protein aggregate present in the sample after incubation consisted of extended filaments with ordered structures. This fibrillar material possesses extensive beta-sheet structure as revealed by far-UV CD and IR spectroscopy. Furthermore, the fibrils exhibit increased Thioflavin T fluorescence.