Identification of α-tubulin alpha-1B chain as a target of asiatic acid using chemical proteomics in HepG2 hepatoma cells.

Asiatic acid (AA) is a naturally occurring pentacyclic triterpene isolated from Centella asiatica and has various biological effects, most notably anticancer effects. While numerous investigations have demonstrated the possible mechanism underlying AA's anticancer action, the precise protein target of AA remains unclear. In this study, the protein target of AA in HepG2 hepatoma cells was identified using the AfBPP-based chemoproteomic approach. Initially, a diazirine and alkyne group modified AA photoaffinity probe was synthesized. Then, using mass spectrometry analysis, 13 putative target proteins were identified with high confidence. Combined with the competition bands in in situ fluorescence scanning, the α-tubulin alpha-1B chain (TUBA1B) was identified as the target protein of AA. Subsequently, the direct interaction between AA and TUBA1B was verified by surface plasmon resonance, pull-down and cellular thermal shift experiments, drug affinity responsive target stability assay, and molecular docking. This research will offer fresh perspectives on how AA prevents liver cancer at the molecular level.