Recognition of a Single β-D-Xylopyranose Molecule by Xylanase GH11 from Thermoanaerobacterium saccharolyticum

Abstract

The endo-β-1,4-xylanase glycosyl hydrolase (GH11) decomposes the backbone of xylan into xylooligosaccharides or xylose. These enzymes are important for industrial applications in the production of biofuel, feed, food, and value-added materials. β-D-xylopyranose (XYP, also known as β-D-xylose) is the fundamental unit of the substrate xylan, and understanding its recognition is fundamental for the initial steps of GH11’s molecular mechanism. However, little is known about the recognition of a single XYP molecule by GH11. In this study, the crystal structures of GH11 from Thermoanaerobacterium saccharolyticum (TsaGH11) complexed with an XYP molecule were determined at a resolution of 1.7–1.9 Å. The XYP molecule binds to subsite −2 of the substrate-binding cleft. The XYP molecule is mainly stabilized by a π–π interaction with the conserved Trp36 residue. The O2 and O3 atoms of XYP are stabilized by hydrogen bond interactions with the hydroxyl groups of Tyr96 and Tyr192. The conformation of the thumb domain of TsaGH11 does not play a critical role in XYP binding, and XYP binding induces a shift in the thumb domain of TsaGH11 toward the XYP molecule. A structural comparison of TsaGH11 with other GH11 xylanases revealed that the XYP molecule forms π–π stacking with the center between the phenyl and indoline ring of Trp36, whereas the XYP molecule unit from xylobiose or xylotetraose forms π–π stacking with the indoline of Trp36, which indicates that the binding modes of the substrate and XYP differ. These structural results provide a greater understanding of the recognition of XYP by the GH11 family.