Structural basis for regulating lipopolysaccharide transmembrane transport.

Abstract

Gram-negative bacteria are surrounded by a multilayered cell envelope with a mostly impermeable outer membrane that provides intrinsic resistance to many antibiotics1-5. The outer membrane is an asymmetrical bilayer with phospholipids in the inner leaflet and lipopolysaccharide (LPS) in the outer leaflet6,7. An LPS transport (Lpt) machine called LptB2FGCADE moves LPS across a protein bridge from the inner membrane to the outer membrane8,9. LPS biosynthesis is regulated to prevent toxic accumulation of LPS molecules in the inner membrane during growth10-12. Whether LPS transport across the Lpt bridge is also regulated has been unclear. Here we present three structures of the trans-envelope Lpt complex in LPS-free, LPS-bound and ATP-bound states, along with a structure of a partial bridge. These structures, combined with biochemical experiments, show that Lpt bridge assembly triggers movement of the transmembrane helix of LptC (TM-LptC) in an LPS-dependent manner, resulting in increased ATP binding and hydrolysis. We also show that LPS transport in vivo requires bridge formation and movement of the TM-LptC. Our data support a model in which assembled Lpt bridges respond to the presence of LPS in the inner membrane to turn on transport by moving the TM-LptC, thus coordinating LPS transport activity with bridge assembly and the presence of LPS at the inner membrane.