The solution structure of PapGII from uropathogenic Escherichia coli and its recognition of glycolipid receptors

Abstract

Uropathogenic Escherichia coli (UPEC) is the primary cause of symptomatic urinary tract infection. The P-pili, a bacterial surface organelle, mediates the bacterial host--cell adhesion. The PapG adhesin has generated much interest in recent years, not only because of its clinical value, i.e. in the prevention of microbial adherence, but also because of its ability to promote virulence. Using multidimensional nuclear magnetic resonance (NMR) and deuteration we have determined the solution structure of the adhesin domain from PapGII (PapGII-198). The novel structure of PapGII-198 is composed of a large elongated jellyroll motif. Despite an automated search of the structural database failing to reveal any similar proteins, PapGII adhesin shares some structural similarities with FimH. Furthermore, interpretation of NMR-titration data has enabled us to identify the putative binding site for the globoseries of oligosaccharides. This work provides insight into UPEC pathogenesis as well as aiding the development of preventative therapies and the guidance of future mutagenesis programmes.

Fig. 1. The solution structure of PapGII-198. (A) Stereo-image backbone traces for the 10 lowest energy structures of PapGII-198 superimposed onto the mean coordinate position. Superimposition is based on the backbone atoms of 1–19, 25–70, 79–92, 96–195. The average structure is shown in orange. (B) Schematic diagram illustrating the arrangement of secondary structure elements for PapGII-198. β-strands are shown as blue arrows, helices as red cylinders and the strand insertion to the jellyroll motif is shaded magenta. (C) Sequence alignment and topology for the PapG family of adhesins. The residues highlighted in red and green represent identity and conservation with the PapG class II sequence, respectively. The approximate location of secondary structure elements is also indicated; helices delineated as tubes and β-strands as arrows. The strand insertion to the jellyroll motif is shaded magenta. The amino acid positions are shown for PapGII from DS17 and correspond to residues 21–219 in full-length PapGII. Asterisks represent amide resonances that are significantly perturbed upon the addition of galabiose.

Fig. 1. The solution structure of PapGII-198. (A) Stereo-image backbone traces for the 10 lowest energy structures of PapGII-198 superimposed onto the mean coordinate position. Superimposition is based on the backbone atoms of 1–19, 25–70, 79–92, 96–195. The average structure is shown in orange. (B) Schematic diagram illustrating the arrangement of secondary structure elements for PapGII-198. β-strands are shown as blue arrows, helices as red cylinders and the strand insertion to the jellyroll motif is shaded magenta. (C) Sequence alignment and topology for the PapG family of adhesins. The residues highlighted in red and green represent identity and conservation with the PapG class II sequence, respectively. The approximate location of secondary structure elements is also indicated; helices delineated as tubes and β-strands as arrows. The strand insertion to the jellyroll motif is shaded magenta. The amino acid positions are shown for PapGII from DS17 and correspond to residues 21–219 in full-length PapGII. Asterisks represent amide resonances that are significantly perturbed upon the addition of galabiose.

Fig. 1. The solution structure of PapGII-198. (A) Stereo-image backbone traces for the 10 lowest energy structures of PapGII-198 superimposed onto the mean coordinate position. Superimposition is based on the backbone atoms of 1–19, 25–70, 79–92, 96–195. The average structure is shown in orange. (B) Schematic diagram illustrating the arrangement of secondary structure elements for PapGII-198. β-strands are shown as blue arrows, helices as red cylinders and the strand insertion to the jellyroll motif is shaded magenta. (C) Sequence alignment and topology for the PapG family of adhesins. The residues highlighted in red and green represent identity and conservation with the PapG class II sequence, respectively. The approximate location of secondary structure elements is also indicated; helices delineated as tubes and β-strands as arrows. The strand insertion to the jellyroll motif is shaded magenta. The amino acid positions are shown for PapGII from DS17 and correspond to residues 21–219 in full-length PapGII. Asterisks represent amide resonances that are significantly perturbed upon the addition of galabiose.

Fig. 2. The structural comparison between PapGII and the FimH adhesins. (A) Ribbon representation of PapGII-198. (B) Ribbon representation of the FimH adhesin domain. β-strands are shown as blue arrows, helices as red cylinders and the strand insertions to the jellyroll motif are shaded magenta.

Fig. 3. Identification of the binding site for galabiose in PapGII-198. (A) Region of the ¹H-¹⁵N HSQC NMR spectrum for ²H,¹³C,¹⁵N-labelled PapGII-198 with the identical region of the spectrum for ²H,¹³C,¹⁵N-labelled PapGII-198 with eqimolar amounts of galabiose overlaid (red). Peaks corresponding to amides that are affected upon addition of galabiose are labelled with their residue number. (B) Column chart of summed ¹H/¹⁵N chemical shift perturbations in the presence of galabiose (weighted n according to ¹H and ¹⁵N chemical shift ranges for PapGII-198 i.e 4.7δ1_H + δ15_N). Contact-surface representation (C) for the structure of PapGII-198. Residues shown in red and yellow experience ¹H/¹⁵N chemical shift perturbations in excess of 1 p.p.m. and between 0.35 and 1 p.p.m., respectively. Only solvent-accessible residues are annotated, as these are most likely to be directly involved in binding.