STRUCTURE AND FUNCTION OF THE XENOBIOTIC SUBSTRATE-BINDING SITE AND LOCATION OF A POTENTIAL NON-SUBSTRATE-BINDING SITE IN A CLASS PI-GLUTATHIONE-S-TRANSFERASE

Complex structures of a naturally occurring variant of human class pi glutathione S-transferase 1-1 (hGSTP1-1) with either S-hexylglutathion e or -glutathionyl)-10-hydroxy-9,10-dihydrophenanthrene [(9R,10R)-GSPh en] have been determined at resolutions of 1.8 and 1.9 Angstrom, respe ctively. The crystal structures reveal that the xenobiotic substrate-b inding site (H-site) is located at a position similar to that observed in class mu GST 1-1 from rat liver (rGSTM1-1). In rGSTM1-1, the H-sit e is a hydrophobic cavity defined by the side chains of Y6, W7, V9, L1 2, I111, Y115, F208, and S209. In hGSTP1-1, the cavity is approximatel y half hydrophobic and half hydrophilic and is defined by the side cha ins of Y7, F8, V10, R13, V104, Y108, N204, and G205 and five water mol ecules. A hydrogen bond network connects the five water molecules and the side chains of R13 and N204. V104 is positioned such that the intr oduction of a methyl group (the result of the V104I mutation) disturbs the H-site water structure and alters the substrate-binding propertie s of the isozyme. The hydroxyl group of Y7 forms a hydrogen bond (3.2 Angstrom) with the sulfur atom of the product. There is a short hydrog en bond (2.5 Angstrom) between Y108 (OH) and (9R, 10R)-GSPhen (O5), in dicating the hydroxyl group of Y108 as an electrophilic participant in the addition of glutathione to epoxides. An N-(2-hydroxethyl)piperazi ne-N'-2-ethanesulfonic acid (HEPES) molecule is found in the cavity be tween beta 2 and alpha I. The location and properties of this HEPES-bi nding site fit a possible non-substrate-binding site that is involved in noncompetitive inhibition of the enzyme.