Thermodynamic origin of fenugreek phytochemical binding to the ASC pyrin domain for inflammation inhibition

Abstract

Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is an essential adaptor protein that regulates inflammasome signaling by binding to NOD-like receptor proteins containing pyrin 3 (NLRP3), crucial for the activation of neuroinflammation. Fenugreek phytochemicals are well-known substances with anti-inflammatory properties. The binding interactions of fenugreek phytochemicals toward the ASC pyrin domain (PYD) protein for inflammation inhibition are largely unknown. To this end, all-atom unbiased molecular dynamics simulations that are a total of 58.33 μs long, including absolute binding free energy and umbrella sampling simulations, are performed for unbound and bound ASC–fenugreek complexes. Our calculations reveal that luteolin, one of the flavonoids in fenugreek, previously known to block NLRP3 inflammasome activation and ASC oligomerization, has the strongest binding to ASC among the major phytochemicals, interacting favorably through hydrogen bonds and driven by enthalpic interactions. Electrostatic interactions primarily govern the bindings of flavonoids and alkaloids, whereas van der Waals interactions dictate the binding of saponins. The higher interface water entropy near the binding residues of the ASC–luteolin complex facilitates the binding of luteolin with ASC. The binding residues of ASC toward luteolin, predicted from hydrogen bond occupancy, native contact analyses, and negative binding enthalpy, are similar to those of NLPR3 known from earlier experiments. This suggests that luteolin binding to ASC can potentially block ASC–NLRP3 binding and activation, which regulate inflammatory ASC oligomerization. The study provides insights for governing the binding interactions of fenugreek phytochemicals towards ASC to inhibit inflammasome complex formation, which controls apoptotic signaling pathways, alongside facilitating the drug discovery process for the future.