Shukdeb Sing, Arindam Das, Suraj Saha, Gouranga Jana, Anirban Basu
Proteins fold abnormally resulting in amyloid aggregates, which cause various degenerative diseases. In this work, the antiamyloidogenic potential of the benzophenanthridine alkaloid, sanguinarine (SNR), has been studied for inhibiting human insulin (HI) fibrillation. Firstly, the interaction of SNR with HI was studied under amyloidogenic conditions using steady-state fluorescence and molecular docking. The intrinsic fluorescence of HI was significantly quenched by SNR and the quenching mechanism was static. The fibrillation kinetics was monitored using a Thioflavin T assay based on fluorescence, which demonstrated that SNR inhibited the growth of HI fibrils markedly. This was further complemented by a Congo red assay based on absorbance. Nile red assays, ANS assays and intrinsic fluorescence studies also supported the fibril arresting ability of SNR. Circular dichroism studies provided evidence that SNR reduced the β-sheet content of the fibrillar species, which in turn implied that fibrillogenesis was suppressed by SNR. AFM imaging revealed that the amount of fibrils generated in the presence of SNR was reduced significantly. Additionally, we found that SNR possesses the ability to defibrillate the preformed fibrillar aggregates. Therefore, the findings from this study indicate that SNR possesses a significant inhibitory effect on HI fibrillation.