The Tyrosine Gate as a Potential Entropic Lever in the Receptor-Binding Site of the Bacterial Adhesin FimH

Abstract

Uropathogenic Escherichia coli (UPEC) are the major causative agents of urinary tract infections. During infection, UPEC adhere to mannosylated glycoreceptors on the urothelium via the FimH adhesin located at the tip of type 1 pili. Synthetic FimH antiadhesives such as alkyl and phenyl α-D-mannopyranosides are thus ideal candidates for the chemical interception of this crucial step in pathogenesis. The crystal structures of the FimH lectin domain in its ligand-free form and in complexes with eight medium- and high-affinity mannopyranoside inhibitors are presented. The thermodynamic profiles of the FimH-inhibitor interactions indicate that the binding of FimH to α-D-mannopyranose is enthalpy-driven and has a negative entropic change. Addition of a hydrophobic aglycon influences the binding enthalpy and can induce a favorable entropic change. The alleviation of the entropic cost is at least in part explained by increased dynamics in the tyrosine gate (Tyr48 and Tyr137) of the FimH receptor-binding site upon binding of the ligand. Ligands with a phenyl group directly linked to the anomeric oxygen of α-D-mannose introduce the largest dynamics into the Tyr48 side chain, because conjugation with the anomeric oxygen of α-D-mannose forces the aromatic aglycon into a conformation that comes into close contact (≈2.65 Å) with Tyr48. A propargyl group in this position predetermines the orientation of the aglycon and significantly decreases affinity. FimH has the highest affinity for α-D-mannopyranosides substituted with hydrophobic aglycons that are compatible in shape and electrostatic properties to the tyrosine gate, such as heptyl α-D-mannose.

The Tyrosine Gate of the Bacterial Lectin FimH: A Conformational Analysis by NMR Spectroscopy and X‐ray Crystallography

“…ChemBioChem 2015, 16,1235 -1246 www.chembiochem.org ferent from that observed in the "pseudo-apo" X-ray structure [26] and in ar ecently solved NMR solution structure [25] (Figure S2). Apart from Tyr48, the conformation of the loop residues is remarkably conserved in all FimH-CRD crystal structures.…”

“…The shifts were larger for aromatic aglycones than for aliphatic ones. In the open conformation seen in the "pseudoapo" crystal structure [26] the Tyr48 ring is distant from the loop residues.I nt he closed conformation,i ti sr otated towards the loop such that Glu50 H N comes closer and within 308 of the ring plane,w here it experiences as trong deshielding aromatic ring current. In contrast, Thr51 H N points almostd irectly into the ring in aT -shaped orientation in whichastrong shielding field causesu pfields hifts.…”

“…[26] The latter is considered a" pseudo-apo" structure, although the bound ethane-1,2-diol could still have an influence on the binding site. Whereas the Tyr137 side chain in these structures remains in the orientation seen in the closed conformation,t he Tyr48 side chain is rotatedt owards the mannose pocket by about 3.…”

“…Wellens et al proposed that the open conformation represents the minimum-energy conformer of FimH and that the closed conformation is only stabilized by favorable interactions with hydrophobic aglycones. [26] This would correspond to ac onformational selection process in which the ligand binds to ac onformer from ap reexisting set of substates. [28] Nevertheless, an induced-fit mechanism also seems possible.…”

“…Unfortunately,i nm any FimH-mannoside co-crystal structures the binding pocket residues or the ligand are involved in crystal lattice contacts potentially affecting the binding pocket geometry. [26][27] Therefore, more experiments are needed to study the structure and dynamics of FimH-antagonist complex formation in solution.…”