Enzymology
Structural and thermodynamic insights into β-1,2-glucooligosaccharide capture by a solute-binding protein in Listeria innocua

https://doi.org/10.1074/jbc.RA117.001536Get rights and content
Under a Creative Commons license
open access

β-1,2-Glucans are bacterial carbohydrates that exist in cyclic or linear forms and play an important role in infections and symbioses involving Gram-negative bacteria. Although several β-1,2-glucan–associated enzymes have been characterized, little is known about how β-1,2-glucan and its shorter oligosaccharides (Sopns) are captured and imported into the bacterial cell. Here, we report the biochemical and structural characteristics of the Sopn-binding protein (SO-BP, Lin1841) associated with the ATP-binding cassette (ABC) transporter from the Gram-positive bacterium Listeria innocua. Calorimetric analysis revealed that SO-BP specifically binds to Sopns with a degree of polymerization of 3 or more, with Kd values in the micromolar range. The crystal structures of SO-BP in an unliganded open form and in closed complexes with tri-, tetra-, and pentaoligosaccharides (Sop3–5) were determined to a maximum resolution of 1.6 Å. The binding site displayed shape complementarity to Sopn, which adopted a zigzag conformation. We noted that water-mediated hydrogen bonds and stacking interactions play a pivotal role in the recognition of Sop3–5 by SO-BP, consistent with its binding thermodynamics. Computational free-energy calculations and a mutational analysis confirmed that interactions with the third glucose moiety of Sopns are significantly responsible for ligand binding. A reduction in unfavorable changes in binding entropy that were in proportion to the lengths of the Sopns was explained by conformational entropy changes. Phylogenetic and sequence analyses indicated that SO-BP ABC transporter homologs, glycoside hydrolases, and other related proteins are co-localized in the genomes of several bacteria. This study may improve our understanding of bacterial β-1,2-glucan metabolism and promote the discovery of unidentified β-1,2-glucan–associated proteins.

ABC transporter
carbohydrate-binding protein
carbohydrate metabolism
X-ray crystallography
molecular dynamics
isothermal titration calorimetry (ITC)
β-1,2-glucan
β-1,2-glucooligosaccharide
solute-binding protein
sophorooligosaccharide
sugar-binding protein

Cited by (0)

This work was supported in part by the Platform Project for Support in Drug Discovery and Life Science Research (Platform for Drug Discovery, Informatics, and Structural Life Science) from the Japan Agency for Medical Research and Development (AMED) and by JSPS-KAKENHI Grants 15H02443 and 26660083 (to S. F.). The authors declare that they have no conflicts of interest with the contents of this article.

The atomic coordinates and structure factors (codes 5YSB, 5YSD, 5YSE, and 5YSF) have been deposited in the Protein Data Bank (http://wwpdb.org/).