We will use X-ray crystallography to investigate the type VII secretion system. Clockwise form top left: Diffraction pattern of a protein crystal, model of the type VII secretion system, protein crystals (µm scale) of a membrane protein complex, cartoon representation of three-dimensional structures of secreted type VII substrate proteins

Pathogenic mycobacteria have evolved sophisticated mechanisms to subvert the human immune defence. Our group aims to understand one of their most important mechanisms - the secretion of proteins by the type VII secretion system.

Tuberculosis is an infectious disease caused by various strains of mycobacteria, which according to recent figures from the World Health Organisation (WHO) accounted for 1.3 million deaths in 2012. Currently about one third of the world’s population carries a latent tuberculosis infection, which makes tuberculosis a major health threat.

Mycobacteria are commonly spread through aerosols such as those generated by coughing. Once inhaled, these pathogens are engulfed in the lungs by alveolar macrophages and targeted for destruction. However, mycobacteria can escape this fate through secretion of specific proteins by a major protein secretion pathway, the ESX/ type VII secretion system, which comprises of five distinct secretion machines, each forming multi-component nano-machines of immense size that are situated in the cell envelope. Each secretion machine has a differential substrate specificity and is believed to be employed at different stages of the infection. For example, ESX-1 secreted proteins enable mycobacteria to escape the phagolysosome in the macrophage, the compartment in which the bacteria are lysed, while ESX-5 secreted proteins trigger granuloma formation. A granuloma is a cluster of immune cells which mycobacteria exploit as replication niches, or for long term persistence until reactivation causes the outbreak of tuberculosis.

Little is known about the structure and the secretion mechanism of the type VII machinery. It has been shown that a set of six conserved proteins EccA-E and MycP are essential for secretion of the type VII substrate. There are four core components EccBCDE which are embedded in the cytoplasmic membrane where they assemble into a stable ~1.5 MDa complex, which forms presumably a secretion pore. The protein transport across the type VII secretion system is powered by one or more ATPases (EccA and EccC) of which EccA is located in the cytosol and may be recruited to the type VII core complex upon substrate binding. Protease MycP processes the transported substrate. Once in the periplasm, the protein substrate is possibly secreted by an independent outer membrane secretion system to the extracellular space.

Due to their unambiguous role in virulence, our research will focus on the ESX-1 and ESX-5 secretion machines as models of type VII secretion systems in pathogenic mycobacteria. Our goal is to determine the first three-dimensional structures of these two type VII secretion machines by state-of-the-art structural methods such as cryo-electron tomography, cryo-electron microscopy and X-ray crystallography, permitting functional studies to generate a model of the type VII secretion mechanism.

Overall, this project will improve our understanding of the pathogen-host communication in tuberculosis and has the potential to provide the structural basis for the development of new therapeutics against tuberculosis.