Structure of the mycobacterial ESX-5 type VII secretion system membrane complex by single-particle analysis

Abstract

Mycobacteria are characterized by their impermeable outer membrane, which is rich in mycolic acids. To transport substrates across this complex cell envelope, mycobacteria rely on type VII (also known as ESX) secretion systems. In Mycobacterium tuberculosis, these ESX systems are essential for growth and full virulence and therefore represent an attractive target for anti-tuberculosis drugs. However, the molecular details underlying type VII secretion are largely unknown, due to a lack of structural information. Here, we report the molecular architecture of the ESX-5 membrane complex from Mycobacterium xenopi determined at 13 Å resolution by electron microscopy. The four core proteins of the ESX-5 complex (EccB, EccC, EccD and EccE) assemble with equimolar stoichiometry into an oligomeric assembly that displays six-fold symmetry. This membrane-associated complex seems to be embedded exclusively in the inner membrane, which indicates that additional components are required to translocate substrates across the mycobacterial outer membrane. Furthermore, the extended cytosolic domains of the EccC ATPase, which interact with secretion effectors, are highly flexible, suggesting an as yet unseen mode of substrate interaction. Comparison of our results with known structures of other bacterial secretion systems demonstrates that the architecture of type VII secretion system is fundamentally different, suggesting an alternative secretion mechanism.

New insights into the mycobacterial PE and PPE proteins provide a framework for future research

“…A promising new approach that could overcome this obstacle is the recent success in expression of a functional ESX‐5 system from M. xenopi in M. smegmatis , which enabled resolution of the first TypeVII secretion system structure by cryo‐electron microscopy. This heterologously expressed ESX‐5 system was able to facilitate secretion of the PPE‐SVP protein PPE18 as well as EsxN in M. smegmatis (Beckham et al , ). No secretion of PE_PGRS proteins has been reported in this model, which is also not expected to occur, since no functional PPE38 is present in this system.…”

“…Together, these exciting and seemingly contradictory findings create a puzzling tension in the field that begs for a unifying explanation which integrates these findings. Together, recent work has provided new roads forward that have the potential to finally bring more insight into the actual function of PE_PGRS and PPE‐MPTR proteins (Ates et al , ; ; Beckham et al , ).…”

“…Similarly, different methods of secretion analysis can come to different results depending on timescale, growth medium and the detection method used. Finally, many studies investigating PE and PPE proteins use Mycobacterium smegmatis as a model organism, which has no functional ESX‐5 system and is therefore unable to solubly express or translocate the majority of PE and PPE proteins (Abdallah et al , ; Sampson, ; Gröschel et al , ; Beckham et al , ).…”

Esx Systems and the Mycobacterial Cell Envelope: What's the Connection?

“…The ESX-5 membrane complex from Mycobacterium xenopi was recently resolved to 13 Å by electron microscopy and revealed a novel oligomeric complex with 6-fold symmetry. The complex resides exclusively within the cytoplasmic membrane and hints at a novel mechanism of protein translocation (145). Higherresolution structures have been solved for individual components in the membrane complex (146)(147)(148)(149)(150).…”

Structural variability of EspG chaperones from mycobacterial ESX-1, ESX-3 and ESX-5 type VII secretion systems

“…Each consists of ATPases, membrane proteins, a protease, accessory proteins, and secreted substrates [6,7]. Four conserved components of the ESX-5 system -EccB 5 , EccC 5 , EccD 5 and EccE 5 -form a platform complex with six-fold symmetry that is embedded in the mycobacterial inner membrane [8,9]. The ESX-1 core complex is composed of paralogous components [10], which suggests that all ESX systems assemble into similar complexes.…”