Fig 1.
Core LPS structure of Salmonella enterica rough mutant strains.
The LPS phenotypes (Ra, Rb1, Rb2, Rb3, Rc, Rd1, Rd2, and Re) and the mutant strains R5 and R7 are shown (modified from Mansfield and Forsythe, 2001 [29]). Glc: glucose; GlcNAc: N-acetyl-glucosamine; Gal: galactose; Hep: L-D-Heptose; P: phosphate; PEtn: phosphoethanolamine; Kdo: 3-deoxy-D-manno-oct-2-ulosonic acid.
Table 1.
Data collection and processing.
Fig 2.
Structure of the product of mild acid hydrolysis of S. enterica Minnesota rough mutants R5 and R7.
Glc: glucose; Hep: L-D-Heptose; Kdo: 3-deoxy-D-manno-oct-2-ulosonic acid.
Fig 3.
Structure of the 4,7 closure furanoid derivative (anhydro Kdo) of Kdo following LPS delipidation by mild acid hydrolysis and β-elimination of the Kdo O4 substituent, reported by Shrive and co-workers (PDB ID 4E52, ligand KD5).
Anhydro Kdo is proposed to be a racemic mixture (Auzanneau et al., 1991 [43]) with the 2-oxobutanoic acid side chain at C4 both alpha and beta to the glycosidic bond (at C5). (a) Original Kdo numbering retained. (b) Numbering according to pdb entry 4E52 (alpha) and used for the structure here.
Fig 4.
Electron density for the Salmonella enterica oligosaccharides bound to rfhSP-D (subunit B) with HepI in the Ca1 binding site.
The calcium ion is in green and the four calcium and ligand coordinating residues Glu321, Asn323, Glu329 and Asn341 are shown in cpk. (a) R7 oligosaccharide KdoI (anhydro)-HepI-HepII (b) R5 oligosaccharide KdoI (anhydro)-HepI-HepII-Glc1. Maps are 2mFo-DFc contoured at 1σ.
Fig 5.
S. enterica R5 oligosaccharide bound to rfhSP-D (subunit C) with the inner core HepI in the Ca1 binding site.
KdoI (anhydro) interacts with both Asp325 and Arg343. Similar binding is observed in subunit B and in subunits B and C of the R7-bound structure which lacks the terminal glucose GlcI. Calcium coordinating bonds are represented by dots and protein-ligand interactions by dashes. The interactions of Glu333 are also indicated.
Table 2.
Calcium and ligand binding distances (Å).
Fig 6.
S. enterica R5 oligosaccharide bound to rfhSP-D (subunit A) with the terminal glucose GlcI in the Ca1 binding site.
HepI interacts with Arg343. No ligand was observed in subunit A of the R7-bound structure due to crystal lattice constraints. Calcium coordinating bonds are represented by dots and protein-ligand interactions by dashes. The interactions of Glu333 are also indicated.
Fig 7.
(a) Superposition of subunit A (GlcI bound) in cyan and subunit B (HepI bound) in yellow of the R5 LPS-bound structure following a least-squares fit of the CRD protein main chain atoms. (b) Superposition of subunit A (GlcI bound) in cyan of the R5 LPS-bound structure and subunit A (Glc1 bound) in coral of the maltose-bound structure [14] following a least-squares fit of the CRD protein main chain atoms.
Fig 8.
Model of complete S. enterica R5 oligosaccharide bound to rfhSP-D with the inner core HepI in the Ca1 binding site.
KdoI (intact) is positioned to interact with Asp325, with KdoII and KdoIII interacting with Arg343 and Glu347 respectively. (a) subunit C, R5-bound structure (b) the rfhSP-D trimer with R5 similarly placed in each subunit and the link to lipid A from KdoI O2' indicated by ★.