Molecular mechanisms of Lobeline-mediated inhibition of lysozyme amyloidogenesis: A synergistic approach using biophysical and cheminformatics techniques.

Numerous pathological conditions, collectively termed amyloidosis, are associated with the aggregation of misfolded proteins under stressed physiochemical conditions. Natural compounds capable of modulating protein aggregation or disassembling preformed fibrils hold promise as potential therapeutic candidates for treating aggregation-related diseases. In this study, we aim to examine the binding interaction and effectiveness of Lobeline (Lob), a piperidine alkaloid, in preventing the formation of acid-denatured Lysozyme (Lyz) amyloid using various spectroscopic, cheminformatics and imaging techniques. Steady-state and time-resolved fluorescence measurements confirm a direct interaction between Lyz and Lob with a binding constant of ∼106 M-1 with a 1:1 binding stoichiometry. The association has been found to be spontaneous and is driven by entropy involving non-electrostatic interactions. Molecular Docking shows that Lob stabilizes Lyz by hydrophobic and hydrophilic interactions. The anti-amyloid properties of Lob in Lyz amyloid fibrils are assessed through a range of in vitro techniques, including turbidity measurement, dynamic light scattering (DLS), Thioflavin T (ThT) fluorescence, Circular Dichroism studies and Field Emission Scanning Electron Microscopy (FESEM) imaging. These studies demonstrate that Lob halts the fibrillation of acid-treated Lyz at the nucleation stage by providing alternative pathways for hydrogen bonding and other weak interactions with key amino acid residues necessary for the formation of oligomers and fibrils.