Gelsemine, a naturally occurring indole alkaloid derived from plants of the Gelsemium species of the Gelsemiaceae family, has been extensively investigated for its neuroprotective and anti-inflammatory properties. Recent studies have demonstrated that gelsemine exerts neuroprotective effects against beta-amyloid (A beta) oligomers, a key neurotoxic peptide implicated in the pathogenesis of Alzheimer's disease (AD). However, despite these beneficial effects, the precise molecular targets underlying gelsemine's neuroprotective actions in AD remain unidentified. Here, we employed a combination of bioinformatic, biochemical, and functional assays in neuronal models to investigate the mechanism of gelsemine's action in AD cellular models. Our findings indicate that gelsemine inhibits the activity of transglutaminase 2 (TG2), an enzyme involved in protein cross-linking with emerging roles in A beta aggregation and neurotoxicity. Molecular modeling and biochemical analyses reveal that gelsemine interacts with the TG2 catalytic site, leading to its inhibition. Furthermore, gelsemine modulates the TG2-mediated A beta aggregation process, thereby attenuating A beta-induced neurotoxicity and preserving neuronal function. These findings establish TG2 as a previously unrecognized molecular target of gelsemine and underscore the potential of Gelsemium-derived alkaloids as neuroprotective agents. The modulation of TG2 activity by natural alkaloids may provide a novel therapeutic approach for mitigating A beta toxicity and preserving neuronal function in AD.