PT  - JOURNAL ARTICLE
AU  - Sean Miletic
AU  - Dirk Fahrenkamp
AU  - Nikolaus Goessweiner-Mohr
AU  - Jiri Wald
AU  - Maurice Pantel
AU  - Oliver Vesper
AU  - Vadim Kotov
AU  - Thomas C. Marlovits
TI  - Substrate-engaged type III secretion system structures reveal gating mechanism for unfolded protein translocation
AID  - 10.1101/2020.12.17.423328
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 2020.12.17.423328
4099  - http://biorxiv.org/content/early/2020/12/18/2020.12.17.423328.short
4100  - http://biorxiv.org/content/early/2020/12/18/2020.12.17.423328.full
AB  - Many bacterial pathogens strictly rely on the activity of type III secretion systems (T3SSs) to secrete and translocate effector proteins in order to establish infection. The central component of T3SSs is the needle complex, a supramolecular machine which assembles a continuous conduit crossing the bacterial envelope and the host cell membrane to allow bacterial effectors to gain entry into the host cell cytoplasm to modulate signal transduction processes. Disruption of this process impairs pathogenicity, providing an avenue for antimicrobial design. However, the molecular principles underlying T3 secretion remain elusive. Here, we report the first structure of an active Salmonella enterica sv. Typhimurium needle complex engaged with the late effector protein SptP in two functional states, revealing the complete 800Å-long secretion conduit and unravelling the critical role of the export apparatus (EA) subcomplex in T3 secretion. Unfolded substrates enter the EA through a hydrophilic constriction formed by SpaQ proteins, which enables side chain-independent transport, explaining heterogeneity and structural disorder of signal sequences in T3SS effector proteins. Above, a methionine gasket formed by SpaP proteins functions as a gate that dilates to accommodate substrates but prevents leaky pore formation to maintain the physical boundaries of compartments separated by a biological membrane. Following gate penetration, a moveable SpaR loop first folds up to then act akin to a linear ratchet to steer substrates through the needle complex. Together, these findings establish the molecular basis for substrate translocation through T3SSs, improving our understanding of bacterial pathogenicity and motility of flagellated bacteria, and paves the way for the development of novel concepts combating bacterial infections.Competing Interest StatementThe authors have declared no competing interest.