Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 111, Issue 13, Pages 4982-4987Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1323516111
Keywords
ABC transporter; antibiotic target; membrane biogenesis
Categories
Funding
- National Science Foundation Graduate Research Fellowship
- Harvard Blavatnik Biomedical Accelerator Fund
- National Institutes of Health [AI081059, GM066174, GM094263]
- Ohio State University
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The cell surface of Gram-negative bacteria contains lipopolysaccharides (LPS), which provide a barrier against the entry of many antibiotics. LPS assembly involves a multiprotein LPS transport (Lpt) complex that spans from the cytoplasm to the outer membrane. In this complex, an unusual ATP-binding cassette transporter is thought to power the extraction of LPS from the outer leaflet of the cytoplasmic membrane and its transport across the cell envelope. We introduce changes into the nucleotide-binding domain, LptB, that inactivate transporter function in vivo. We characterize these residues using biochemical experiments combined with high-resolution crystal structures of LptB pre- and post-ATP hydrolysis and suggest a role for an active site residue in phosphate exit. We also identify a conserved residue that is not required for ATPase activity but is essential for interaction with the transmembrane components. Our studies establish the essentiality of ATP hydrolysis by LptB to power LPS transport in cells and suggest strategies to inhibit transporter function away from the LptB active site.
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