Deciphering the Role of MCE-domain containing Proteins in Outer Membrane Lipid Transport in Gram-negative Bacteria University of Birmingham in United Kingdom

Project Overview

The growing threat of antibiotic-resistant bacteria highlights an urgent need for novel therapeutic strategies. This challenge is particularly acute for Gram-negative bacteria, which are more resilient than their Gram-positive counterparts due to an additional outer membrane. This outer membrane not only protects against antibiotics and detergents but also plays a critical role in pathogenesis, virulence, and nutrient uptake. The complex lipid bilayer of the outer membrane, featuring lipopolysaccharides (LPS) on the outer surface and phospholipids on the inner layer, forms an effective barrier to both hydrophilic and hydrophobic molecules, contributing to their robust defence mechanisms.

Research Focus

This PhD project aims to deepen our understanding of outer membrane biogenesis by investigating the transport of phospholipids from the inner to the outer membrane—a process that remains poorly understood despite decades of research. Recent findings have identified a group of proteins containing Mammalian Cell Entry (MCE) domains as key players in this lipid transport process. These proteins, first identified in Mycobacteria, are widely conserved across bacterial species.

In E. coli, the maintenance of lipid asymmetry (Mla) pathway, particularly MlaD, plays a key role in removing mis-localised phospholipids from the outer membrane, thereby preserving membrane integrity. Two other MCE-domain-containing proteins, PqiB and LetB, have also been implicated in phospholipid transport between membranes, encoding proteins that form a hydrophobic tunnel potentially facilitating lipid transfer across the periplasmic space.

Key Objectives

The primary aim of this project is to unravel the mechanisms by which MCE-domain proteins, such as PqiB and LetB, mediate phospholipid transport in Gram-negative bacteria. Specific objectives include:

• Elucidate what drives activity to address key unknowns surrounding energy requirements, lipid selectivity and regulation.

• Elucidate the mechanistic details of transport to enable understanding of how lipids are extracted and transported between membranes.

• Resolve cellular importance, what is the rationale for requiring multiple systems? Do they transport subsets of lipids?

Methodology

This project will leverage a multidisciplinary approach, combining structural biology, biophysics, and microbiology. Techniques such as cryo-EM, neutron reflectometry, and lipid transport assays will be used to gain a comprehensive understanding of the mechanisms underlying outer membrane biogenesis.

Ideal Candidate

We are looking for a highly motivated and enthusiastic candidate with a background in molecular biology, microbiology, biochemistry, or structural biology. Previous experience with structural techniques or interest in antimicrobial research would be advantageous.

Why Apply?

Joining this project offers a unique opportunity to contribute to cutting-edge research in bacterial cell biology, with the potential to uncover novel targets for therapeutic intervention. You will gain hands-on experience with state-of-the-art techniques and work within a collaborative research environment.

Funding notes:

1. Competition based funding available through the Midlands Integrative Biosciences Training Partnership - https://warwick.ac.uk/fac/cross_fac/mibtp/

https://www.birmingham.ac.uk/research/activity/mibtp

2. Competition based funding available through the Darwin Trust of Edinburgh

3. Self-funded

Please contact [email protected] for more information regarding funding available.

References:

Cooper, B.F., Ratkeviciute, G., Clifton, L.A., Johnston, H., Holyfield, R., Hardy, D.J., Caulton, S.G., Chatterton, W., Sridhar, P., Wotherspoon, P., Hughes, G.W., Hall, S.C., Lovering, A.L. and Knowles, T.J. (2024). "An octameric PqiC toroid stabilises the outer-membrane interaction of the PqiABC transport system." EMBO Rep 25(1): 82-101.

Wotherspoon, P., Johnston, H., Hardy, D.J., Holyfield, R., Bui, S., Ratkeviciute, G., Sridhar, P., Colburn, J., Wilson, C.B., Colyer, A., Cooper, B.F., Bryant, J.A., Hughes, G.W., Stansfeld, P.J., Bergeron, J.R.C. and Knowles, T.J. (2024). "Structure of the MlaC-MlaD complex reveals molecular basis of periplasmic phospholipid transport." Nat Commun 15(1): 6394.