Homology Modeling of Human Uridine-5′-diphosphate-glucuronosyltransferase 1A6 Reveals Insights into Factors Influencing Substrate and Cosubstrate Binding

Abstract

The elimination of numerous endogenous compounds and xenobiotics via glucuronidation by uridine-5′-diphosphate glycosyltransferase enzymes (UGTs) is an essential process of the body’s chemical defense system. UGTs have distinct but overlapping substrate preferences, but the molecular basis for their substrate specificity remains poorly understood. Three-dimensional protein structures can greatly enhance our understanding of the interactions between enzymes and their substrates, but because of the inherent difficulties in purifying and crystallizing integral endoplasmic reticulum membrane proteins, no complete mammalian UGT structure has yet been produced. To address this problem, we have created a homology model of UGT1A6 using I-TASSER to explore, in detail, the interactions of human UGT1A6 with its substrates. Ligands were docked into our model in the presence of the cosubstrate uridine-5′-diphosphate-glucuronic acid, interacting residues were examined, and poses were compared to those cocrystallized with various plant and bacterial glycosyltransferases (GTs). Our model structurally resembles other GTs, and docking experiments replicated many of the expected UGT-substrate interactions. Some bias toward the template structures’ protein–substrate interactions and binding preferences was evident.

A potential implication of UDP-glucuronosyltransferase 2B10 in the detoxification of drugs used in pediatric hematopoietic stem cell transplantation setting: an in silico investigation

“…However, it is also worth noting that there is an experimental evidence indicating the importance of H34 in catalyzing O-glucuronidation reaction, that concluded the need for further investigations to elucidate its function [ 70 ]. It is also likely that binding affinities of the putative ligands may be biased by the template substrate specificities used for homology modelling as it was seen in case of UGT1A6 [ 71 ]. Since UGTs also participates in glucosidation, residues offering specificity for glucuronidation or glucosidation in N-terminal domain may also be biased by the template used for homology modelling(e.g.…”

“…glucuronidation specificity offered by Arg259or glucosidation by UGT2B7) [ 72 ]. However homology modelling successfully implemented to test substrates and inhibitors of other UGT enzymes in the past with a satisfactory level of sensitivity and specificity [ 71 ].…”

Arginine-259 of UGT2B7 Confers UDP-Sugar Selectivity

“…Consistent with this observation, all members of the UGT 1 and 2 families utilise UDP-GlcUA as a cofactor and hence form glucuronide conjugates. The recent UGT1A6 model reported by Smith et al (2020) showed the presence of Arg256 (equivalent to Arg259 in UGT2B7) in the cofactor binding site (Supplementary Figure S1). However, this residue interacted with the phosphate group of UDP-GlcUA.…”

“…Only two MD simulation studies that investigated ligand binding to UGT proteins (viz. UGT1A6 and UGT1A9/10) have been reported to date (Tripathi et al 2015;Smith et al 2020). Neither of these studies provided experimental verification of residues predicted to be involved in UDP-sugar binding, nor insights into the residues that are essential for cofactor selectivity.…”

“…However, this residue interacted with the phosphate group of UDP-GlcUA. Notably, the simulations by Smith et al (2020) were performed for 50ns and did not reach equilibrium, as the rmsd increased with time. In contrast, our simulations show that the rmsd of the C α atoms of UGT2B7 in the presence of UDP-Glc (~2.2 Å) and UDP-GlcUA (~2 Å) stabilised after approximately 50 and 100ns, respectively (Supplementary Figure S2), suggesting longer simulation times are essential for attaining equilibrium.…”