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An Escherichia coli LptA Mutant Defective in Lipopolysaccharide Export
Author(s) -
Ma Bing,
Raetz Christian R.H.
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.815.1
Subject(s) - periplasmic space , bacterial outer membrane , inner membrane , cytosol , lipopolysaccharide , lipid a , biochemistry , mutant , chemistry , escherichia coli , microbiology and biotechnology , biology , membrane , enzyme , gene , endocrinology
Lipopolysaccharide precursors and phospholipids in Escherichia coli are synthesized on the inner surface of the inner membrane and are transported through the periplasm to the outer surface of the outer membrane by a mechanism that is poorly characterized. Some evidence suggests that a periplasmic protein LptA, a cytosolic protein LptB and a membrane‐bound protein LptC are involved in lipopolysaccharide export, but their roles are not clearly defined. We now report a temperature‐sensitive mutant, MB1, with Ser 62 → Cys and Gln 111 → Pro substitutions in LptA. As shown by 32 P‐ and 35 S‐labeling, export of phospholipids and newly synthesized proteins in MB1 were not impaired, but the lipopolysaccharide export to the outer membrane is largely inhibited after 30 min at 42 ° C. Using the lipid A 1‐phosphatase LpxE as the periplasmic marker and the lipid A 3‐O‐deacylase PagL as the outer membrane marker, we were able to determine that the lipid A in MB1 at 42 ° C was terminally localized to the periplasm because it was dephosphorylated by LpxE but was not deacylated by PagL. Moreover, transmission electron microscopy showed that dense particles accumulated at the periplasmic space in MB1 at 42 ° C, consistent with a key role for LptA in lipopolysaccharide trafficking at a step subsequent to the inner membrane flip‐flop but prior to the outer membrane flip‐flop. This research was supported by National Institutes of Health Grant GM‐51310 to C. R. H. Raetz.