Loop Evolutionary Patterns Shape Catalytic Efficiency of TRI101/201 for Trichothecenes: Insights into Protein-Substrate Interactions.

Abstract

Trichothecenes are highly toxic mycotoxins produced by Fusarium fungi, while TRI101/201 family enzymes play a crucial role in detoxification through acetylation. Studies on the substrate specificity and catalytic kinetics of TRI101/201 have revealed distinct kinetic characteristics, with significant differences observed in catalytic efficiency toward deoxynivalenol, while the catalytic efficiency for T-2 toxin remains relatively consistent. In this study, we used structural bioinformatics analysis and a molecular dynamics simulation workflow to investigate the mechanism underlying the differential catalytic activity of TRI101/201. The findings revealed that the binding stability between trichothecenes and TRI101/201 hinges primarily on a hydrophobic cage structure within the binding site. An intrinsic disordered loop, termed loop cover, defined the evolutionary patterns of the TRI101/201 protein family that are categorized into four subfamilies (V1/V2/V3/M). Furthermore, the unique loop displayed different conformations among these subfamilies' structures, which served to disrupt (V1/V2/V3) or reinforce (M) the hydrophobic cages. The disrupted cages enhanced the water exposure of the hydrophilic moieties of substrates like deoxynivalenol and thereby hindered their binding to the catalytic sites of V-type enzymes. In contrast, this water exposure does not affect substrates like T-2 toxin, which have more hydrophobic substituents, resulting in a comparable catalytic efficiency of both V- and M-type enzymes. Overall, our studies provide theoretical support for understanding the catalytic mechanism of TRI101/201, which shows how an intrinsic disordered loop could impact the protein-ligand binding and suggests a direction for rational protein design in the future.