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J. Biol. Chem., Vol. 280, Issue 30, 27679-27687, July 29, 2005

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Loss of Outer Membrane Proteins without Inhibition of Lipid Export in an Escherichia coli YaeT Mutant^*

William T. Doerrler and Christian R. H. Raetz

From the Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710

Escherichia coli yaeT encodes an essential, conserved outer membrane (OM) protein that is an ortholog of Neisseria meningitidis Omp85. Conflicting data with N. meningitidis indicate that Omp85 functions either in assembly of OM proteins or in export of OM lipids. The role of YaeT in E. coli was investigated with a new temperature-sensitive mutant harboring nine amino acid substitutions. The mutant stops growing after 60 min at 44 °C. After 30 min at 44 °C, incorporation of [³⁵S]methionine into newly synthesized OM proteins is selectively inhibited. Synthesis and export of OM phospholipids and lipopolysaccharide are not impaired. OM protein levels are low, even at 30 °C, and the buoyant density of the OM is correspondingly lower. By Western blotting, we show that levels of the major OM protein OmpA are lower in the mutant in whole cells, membranes, and the growth medium. SecA functions as a multicopy suppressor of the temperature-sensitive phenotype and partially restores OM proteins. Our data are consistent with a critical role for YaeT in OM protein assembly in E. coli.

Received for publication, May 2, 2005 , and in revised form, June 2, 2005.

^* This work was supported by National Institutes of Health Grants GM-51310 (to C. R. H. R.) and 1 F32 AI-10613-01 (to W. T. D.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Present address: Dept. of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803.

To whom correspondence should be addressed: Dept. of Biochemistry, Duke University Medical Center, P. O. Box 3711, Durham, NC 27710. Tel.: 919-684-5326; Fax: 919-684-8885; E-mail: raetz{at}biochem.duke.edu.

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