Biophysical and computational insights into the activation mechanism of α-glucosidase by aromatic organophosphate esters.

Organophosphate esters (OPEs) are emerging environmental pollutants with potential estrogenic activity. However, long-term OPE exposure has also been linked to an increased risk of type 2 diabetes mellitus (T2DM), though the molecular mechanisms remain unclear. α-glucosidase plays a critical role in carbohydrate digestion, making it a key target for T2DM. In this study, two typical aromatic organophosphate esters (AOPEs), triphenyl phosphate (TPHP) and tricresyl phosphate (TCP), were selected to investigate how they affect the structure and function of α-glucosidase. Enzyme activity assays revealed that both TPHP and TCP activated α-glucosidase. TPHP exhibited a clear dose-dependent activation, whereas TCP showed no such concentration dependency. The formation of both complexes was driven by hydrophobic interactions. TCP exhibited a higher binding affinity than TPHP, which may be attributed to the distinct structural and electronic properties revealed by density functional theory (DFT) calculations. Computational simulations confirmed that both compounds form stable complexes with α-glucosidase, inducing only subtle local perturbations without significantly altering the overall enzyme fold. This conclusion is supported by stable Rg values, minor CD spectral changes, and a single energy minimum observed in FEL analysis. Furthermore, alanine scanning mutagenesis identified PHE157 and PHE177 as common critical residues for TPHP and TCP binding to α-glucosidase, with energy contributions exceeding 1.5 kcal/mol. Together, these findings provide molecular-level insights into how TPHP and TCP affect the structure and function of α-glucosidase, offering a potential mechanistic explanation for their glucose metabolism-disrupting effects.