Bacterial Cell Envelope Biogenesis

Most bacteria have a size of 1/1000 of a millimetre and propagate as single-cell organism, often with a generation time that can be as short as 20 minutes. Bacteria are also remarkably robust and can survive in a range of different environments. Whilst most bacteria have advantageous roles for humans or the earth's ecosystems, some can cause severe infectious diseases that need to be treated by antibacterial drugs. However, many pathogenic bacteria have acquired the ability to resist antibiotics which became a major burden to our society and makes the development of new antibiotics a high priority. Particularly problematic are pathogenic Gram-negative bacteria. The defining feature of Gram-negative bacteria is their outer membrane, which is intrinsically impermeable to many antibiotics and thus confers a natural resistance to these drugs.

The outer membrane is tightly connected to the cell wall, also called peptidoglycan, which protects the cell from bursting by its internal osmotic pressure. Both, the outer membrane and the cell wall are essential cell envelope structures and their destabilization by chemicals or enzymes can render a bacterial cell susceptible to antibiotics or lead to cell death. Indeed, some of our best antibiotics, like penicillins or glycopepides, directly target the synthesis of the cell wall and few recently identified antibacterial compounds inhibit the biogenesis of the outer membrane.

Growing and dividing bacteria enlarge their cell envelope by synthesizing its components inside the cell and inserting them into the existing layers. Because the outer membrane surrounds the cell wall, the newly synthesized outer membrane components must be transported through the cell wall to the outer membrane. How the cell transports the outer membrane components through the cell wall and how it coordinates outer membrane expansion with cell wall growth to maintain the stability of the cell envelope, is not known. The topic of this project is to decipher the molecular mechanisms of this coordination. We will use molecular biology, biochemical and biophysical methods to determine how the cell wall affects the biogenesis of the outer membrane, and how the dynamic cell envelope processes affect the cell wall structure. In addition, we will investigate how the tight connections between the outer membrane and cell wall affect the cell envelope transport processes for outer membrane growth. We expect that the project will unravel molecular mechanisms that enhance our fundamental knowledge of a bacterial cell and can be targeted by novel antibacterial drugs.

Diderm bacteria have a tripartite cell envelope that contains two membranes, the cytoplasmic membrane and the outer membrane, which contain the periplasm with its stress-bearing net-like peptidoglycan layer. The peptidoglycan layer protects the cell from bursting due to its turgor, whilst the outer membrane with its lipopolysaccharide in the outer leaflet and the porin channels, provides a permeability barrier for many antimicrobial compounds. The outer membrane connects to the peptidoglycan layer by highly abundant proteins and this tight connection ensure the stability of the cell envelope and maintains the barrier function of the outer membrane. Growing and dividing cells expand their cell envelope using macromolecular machineries. The elongasome and divisome complexes enlarge the peptidoglycan layer while trans-envelope export machines transport outer membrane components (lipopolysaccharide, outer membrane proteins and phospholipids) through the periplasm and insert these into the outer membrane. How outer membrane biogenesis and peptidoglycan growth are coordinated is not known. This proposal aims to unravel the molecular mechanisms by which outer membrane biogenesis is coordinated with peptidoglycan growth during normal growth and under stress conditions. We will investigate how the structure of the peptidoglycan affects the functioning of the machine for outer membrane protein assembly. We will study how the peptidoglycan is remodelled during the assembly and disassembly of trans-envelope machines for outer membrane biogenesis. Finally, we will decipher how the dynamics of connections between outer membrane anchored proteins and peptidoglycan affects the biogenesis of the outer membrane and the stability of the cell envelope. Overall, we expect to discover novel mechanisms by which bacteria maintain a stable cell envelope during growth and under stress conditions. Such mechanisms could be targeted by novel drugs to target antimicrobial drug resistance.