Elucidating the mechanisms of outer membrane biogenesis in Gram-negative bacteria

Antibiotic resistance poses a major risk globally. Resistance in Gram-negative bacteria poses the biggest threat due to their complex double-membrane architecture. In particular, the outer membrane provides an impermeable barrier, preventing the uptake of molecules including antibiotics. Thus, the biogenesis of the outer membrane has been a key focus in the fight against antibiotic resistance.
The outer membrane of Gram-negative bacteria is highly asymmetric, with LPS on the outer leaflet and phospholipids in the inner leaflet. The transport of LPS has been widely studied, however the transport of phospholipids is less well understood. Changes in the asymmetry of the outer membrane have been shown to increase the permeability of bacteria and drug sensitivity. Several transport machineries have been noted for their role in phospholipid transport: the Mla, PqiABC, and YebST systems. While diverse in their architecture and arrangement in the periplasm, all these systems contain mammalian cell entry (MCE) domains. However, how MCE domains facilitate lipid transport remains to be elucidated.
This project will mainly focus on PqiABC, with the aim to prove firstly if the system is a phospholipid transporter and uncover the mechanisms behind transport, including the role of the MCE domains. This will use a multi-disciplinary approach of biophysics combined with molecular biology, with a large focus of structural biology. Techniques including electron microscopy, X-ray crystallography, neutron reflectometry and NMR will be utilised to answer the many questions surrounding the function of PqiABC and the mechanistic pathway of transport.