Deciphering the conformational and functional alterations of α-glucosidase induced by emerging tire-derived contaminants 6PPD and 6PPD-quinone.
N-(1, 3-dimethylbutyl)-N'-phenyl-p-phenylenediamine (6PPD) and its transformation product 6PPD-quinone are emerging tire-derived contaminants with disruptive effects on glucose metabolism. Given that α-glucosidase is key to blood glucose homeostasis, this study used multivariate methods to explore how 6PPD and 6PPD-quinone alter its conformation and function. The results show that the ligands spontaneously bind to α-glucosidase via hydrophobic forces, with binding constants (Ka) of 7.312 × 105 M-1 and 5.214 × 104 M-1 at 310 K for 6PPD and 6PPD-quinone, respectively. However, they exert opposing functional effects: 6PPD enhances enzyme activity, whereas 6PPD-quinone exerted an inhibitory effect. Computational alanine scanning mutagenesis (ASM) revealed distinct binding interactions between 6PPD/6PPD-quinone and the catalytic triad of α-glucosidase (Asp214, Glu276, and Asp349), which may partly explain their opposing regulatory effects on α-glucosidase activity. Additionally, conformational analysis indicated that both compounds induce a more compact and stable conformation of α-glucosidase, likely attributed to an increase in α-helix content. Specifically, the α-helix content rose from 25.3 ± 0.2% to 29.7 ± 0.4% in the 6PPD group, and to 27.2 ± 0.3% in the 6PPD-quinone group. Free energy landscape (FEL) analysis further demonstrated that the presence of both compounds shifted the system from two dominant conformations to a single dominant state. Collectively, these findings reveal distinct mechanisms by which tire-derived contaminants modulate α-glucosidase, providing insights into their potential to disrupt glucose homeostasis.