Regulation of Protein Disaggregation and its Cytotoxicity by the Hydrophobic Chain Length of Ammonium-Based Ionic Liquids
Abstract
Amyloid formation and protein aggregation are essential mechanisms in a number of physiologic and pathological circumstances. This study explores the influence of ammonium-based ionic liquids with varying hydrophobic alkyl chain lengths (N2, N4, N6, and N8) on the aggregation and disaggregation of Bone Morphogenetic Protein-2 (BMP-2). Heat-induced aggregation led to a substantial increase in hydrodynamic radius, with measurements revealing an increase from 12 ± 5 nm (native BMP-2) to 400 ± 30 nm. Treatment with N8 further increased the aggregate size to 635 ± 43 nm, supporting the formation of mature fibrils. Conversely, shorter-chain ionic liquids (N2 and N4) reduced the aggregate size significantly to 42 ± 5 nm and 37 ± 11 nm, respectively, indicating effective disaggregation. Spectroscopic analysis confirmed enhanced β-sheet content in fibrillar structures, particularly in N8-treated samples, while FTIR and microscopy revealed structural differences across treatments. Cytotoxicity assays demonstrated that fibrillar aggregates exhibited higher oxidative stress and cell damage, whereas disaggregated structures formed by shorter-chain ionic liquids showed reduced toxicity. These findings underscore the potential of ionic liquids in modulating protein aggregation pathways, offering valuable insights for amyloid research and therapeutic strategies.