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Trapped lipopolysaccharide and LptD intermediates reveal lipopolysaccharide translocation steps across the Escherichia coli outer membrane.

Li X, Gu Y, Dong H, Wang W, Dong C - Sci Rep (2015)

Bottom Line: LptD/E complex forms a N-terminal LPS transport slide, a hydrophobic intramembrane hole and the hydrophilic channel of the barrel, for LPS transport, lipid A insertion and core oligosaccharide and O-antigen polysaccharide translocation, respectively.However, there is no direct evidence to confirm that LptD/E transports LPS from the periplasm to the external leaflet of the outer membrane.By replacing LptD residues with an unnatural amino acid p-benzoyl-L-phenyalanine (pBPA) and UV-photo-cross-linking in E.coli, the translocon and LPS intermediates were obtained at the N-terminal domain, the intramembrane hole, the lumenal gate, the lumen of LptD channel, and the extracellular loop 1 and 4, providing the first direct evidence and "snapshots" to reveal LPS translocation steps across the outer membrane.

View Article: PubMed Central - PubMed

Affiliation: Biomedical Research Centre, Norwich Medical School, University of East Anglia, Norwich Research Park, Norwich, NR4 7TJ, UK.

ABSTRACT
Lipopolysaccharide (LPS) is a main component of the outer membrane of Gram-negative bacteria, which is essential for the vitality of most Gram-negative bacteria and plays a critical role for drug resistance. LptD/E complex forms a N-terminal LPS transport slide, a hydrophobic intramembrane hole and the hydrophilic channel of the barrel, for LPS transport, lipid A insertion and core oligosaccharide and O-antigen polysaccharide translocation, respectively. However, there is no direct evidence to confirm that LptD/E transports LPS from the periplasm to the external leaflet of the outer membrane. By replacing LptD residues with an unnatural amino acid p-benzoyl-L-phenyalanine (pBPA) and UV-photo-cross-linking in E.coli, the translocon and LPS intermediates were obtained at the N-terminal domain, the intramembrane hole, the lumenal gate, the lumen of LptD channel, and the extracellular loop 1 and 4, providing the first direct evidence and "snapshots" to reveal LPS translocation steps across the outer membrane.

No MeSH data available.


Related in: MedlinePlus

Lipopolysaccharide and its outer membrane translocon LptD/E.a lipopolysaccharide of S. typhimurium strain LT2. n = 4–40. LPS contains lipid A, core oligosaccharide and O-antigen. b LptD/E complex structure of S. typhimurium strain LT2. The N-terminal domain of Salmonella LptD is generated by modeling, based on the structure of Shigella flexneri LptD. The positions of residues are detected the cross-linking with LPS are shown in magentas, otherwise in yellow. The residues are selected at the N-terminal domain, the hydrophobic intramembrane hole, the lumenal gate and the lumen of barrel.
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f1: Lipopolysaccharide and its outer membrane translocon LptD/E.a lipopolysaccharide of S. typhimurium strain LT2. n = 4–40. LPS contains lipid A, core oligosaccharide and O-antigen. b LptD/E complex structure of S. typhimurium strain LT2. The N-terminal domain of Salmonella LptD is generated by modeling, based on the structure of Shigella flexneri LptD. The positions of residues are detected the cross-linking with LPS are shown in magentas, otherwise in yellow. The residues are selected at the N-terminal domain, the hydrophobic intramembrane hole, the lumenal gate and the lumen of barrel.

Mentions: All Gram-negative bacteria have an asymmetric bilayer outer membrane with lipopolysaccharide (LPS) forming the outer leaflet and phospholipid composing the inner leaflet12, respectively. Brought together by cations, lipopolysaccharide molecules form a permanent barrier, which is essential for the vitality of most Gram-negative bacteria and plays an important role for drug resistance3. LPS is an amphipathic macromolecule, normally containing three components, lipid A, core oligosaccharide and polysaccharide called O-antigen (Fig. 1a). Lipid A linked to its core oligosaccharide and the bactoprenyl-O-antigen units are each synthesized separately in the cytoplasm and transported by MsbA and Wzx, respectively, to the periplasmic side of the inner membrane, where the O-antigen is then polymerized and ligated en bloc to the lipid A core oligosaccharide terminal end, thus forming mature LPS molecules1.


Trapped lipopolysaccharide and LptD intermediates reveal lipopolysaccharide translocation steps across the Escherichia coli outer membrane.

Li X, Gu Y, Dong H, Wang W, Dong C - Sci Rep (2015)

Lipopolysaccharide and its outer membrane translocon LptD/E.a lipopolysaccharide of S. typhimurium strain LT2. n = 4–40. LPS contains lipid A, core oligosaccharide and O-antigen. b LptD/E complex structure of S. typhimurium strain LT2. The N-terminal domain of Salmonella LptD is generated by modeling, based on the structure of Shigella flexneri LptD. The positions of residues are detected the cross-linking with LPS are shown in magentas, otherwise in yellow. The residues are selected at the N-terminal domain, the hydrophobic intramembrane hole, the lumenal gate and the lumen of barrel.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493717&req=5

f1: Lipopolysaccharide and its outer membrane translocon LptD/E.a lipopolysaccharide of S. typhimurium strain LT2. n = 4–40. LPS contains lipid A, core oligosaccharide and O-antigen. b LptD/E complex structure of S. typhimurium strain LT2. The N-terminal domain of Salmonella LptD is generated by modeling, based on the structure of Shigella flexneri LptD. The positions of residues are detected the cross-linking with LPS are shown in magentas, otherwise in yellow. The residues are selected at the N-terminal domain, the hydrophobic intramembrane hole, the lumenal gate and the lumen of barrel.
Mentions: All Gram-negative bacteria have an asymmetric bilayer outer membrane with lipopolysaccharide (LPS) forming the outer leaflet and phospholipid composing the inner leaflet12, respectively. Brought together by cations, lipopolysaccharide molecules form a permanent barrier, which is essential for the vitality of most Gram-negative bacteria and plays an important role for drug resistance3. LPS is an amphipathic macromolecule, normally containing three components, lipid A, core oligosaccharide and polysaccharide called O-antigen (Fig. 1a). Lipid A linked to its core oligosaccharide and the bactoprenyl-O-antigen units are each synthesized separately in the cytoplasm and transported by MsbA and Wzx, respectively, to the periplasmic side of the inner membrane, where the O-antigen is then polymerized and ligated en bloc to the lipid A core oligosaccharide terminal end, thus forming mature LPS molecules1.

Bottom Line: LptD/E complex forms a N-terminal LPS transport slide, a hydrophobic intramembrane hole and the hydrophilic channel of the barrel, for LPS transport, lipid A insertion and core oligosaccharide and O-antigen polysaccharide translocation, respectively.However, there is no direct evidence to confirm that LptD/E transports LPS from the periplasm to the external leaflet of the outer membrane.By replacing LptD residues with an unnatural amino acid p-benzoyl-L-phenyalanine (pBPA) and UV-photo-cross-linking in E.coli, the translocon and LPS intermediates were obtained at the N-terminal domain, the intramembrane hole, the lumenal gate, the lumen of LptD channel, and the extracellular loop 1 and 4, providing the first direct evidence and "snapshots" to reveal LPS translocation steps across the outer membrane.

View Article: PubMed Central - PubMed

Affiliation: Biomedical Research Centre, Norwich Medical School, University of East Anglia, Norwich Research Park, Norwich, NR4 7TJ, UK.

ABSTRACT
Lipopolysaccharide (LPS) is a main component of the outer membrane of Gram-negative bacteria, which is essential for the vitality of most Gram-negative bacteria and plays a critical role for drug resistance. LptD/E complex forms a N-terminal LPS transport slide, a hydrophobic intramembrane hole and the hydrophilic channel of the barrel, for LPS transport, lipid A insertion and core oligosaccharide and O-antigen polysaccharide translocation, respectively. However, there is no direct evidence to confirm that LptD/E transports LPS from the periplasm to the external leaflet of the outer membrane. By replacing LptD residues with an unnatural amino acid p-benzoyl-L-phenyalanine (pBPA) and UV-photo-cross-linking in E.coli, the translocon and LPS intermediates were obtained at the N-terminal domain, the intramembrane hole, the lumenal gate, the lumen of LptD channel, and the extracellular loop 1 and 4, providing the first direct evidence and "snapshots" to reveal LPS translocation steps across the outer membrane.

No MeSH data available.


Related in: MedlinePlus