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Proc Natl Acad Sci U S A. 2008 Sep 16;105(37):13823-8. doi: 10.1073/pnas.0807028105. Epub 2008 Sep 3.

Periplasmic orientation of nascent lipid A in the inner membrane of an Escherichia coli LptA mutant.

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  • 1Department of Biochemistry, Duke University Medical Center, Box 3711 DUMC, Durham, NC 27710, USA.

Abstract

The core-lipid A domain of Escherichia coli lipopolysaccharide (LPS) is synthesized on the inner surface of the inner membrane (IM) and flipped to its outer surface by the ABC transporter MsbA. Recent studies with deletion mutants implicate the periplasmic protein LptA, the cytosolic protein LptB, and the IM proteins LptC, LptF, and LptG in the subsequent transport of nascent LPS to the outer membrane (OM), where the LptD/LptE complex flips LPS to the outer surface. We have isolated a temperature-sensitive mutant (MB1) harboring the S22C and Q111P substitutions in LptA. MB1 stops growing after 30 min at 42 degrees C. (32)P(i) and [(35)S]methionine labeling show that export of newly synthesized phospholipids and proteins is not severely impaired, but export of LPS is defective. Using the lipid A 1-phosphatase LpxE as a periplasmic IM marker and the lipid A 3-O-deacylase PagL as an OM marker, we show that core-lipid A reaches the periplasmic side of the IM at 42 degrees C in MB1 but not the outer surface of the OM. Electron microscopy of MB1 reveals dense periplasmic material and a smooth OM at 42 degrees C, consistent with a role for LptA in shuttling LPS across the periplasm.

PMID:
18768814
[PubMed - indexed for MEDLINE]
PMCID:
PMC2544538
Free PMC Article

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