Wspólnotowy Serwis Informacyjny Badan i Rozwoju - CORDIS


FUTURE T3SS Streszczenie raportu

Project ID: 311374
Źródło dofinansowania: FP7-IDEAS-ERC
Kraj: Germany

Mid-Term Report Summary - FUTURE T3SS (Bacterial effector secretion: Function and Architecture of the Type 3 Secretion System)

I am very grateful that research in my lab is supported by the ERC. The ERC starting grant allowed my coworkers and me to address several challenging questions in the field of host pathogen interactions during the last 2.5 years.

A recent major breakthrough in our lab was the blocking of the effector transport through the type 3 secretion system(T3SS) using protein chimeras of individual effectors fused to a knotted protein. Locking the T3SS of Shigella flexneri in an actively secreting state by expressing such effector chimeras was crucial for the structural characterization of T3SS intermediates. Based on these findings we started the 3-dimensional structural analysis of T3SS intermediates using cryo-EM. We used a similar strategy to block the protein translocation in other transport systems. Indeed, expressing substrate molecules fused to a knotted protein allowed us to block different bacterial transport systems.

We solved the structure of the Salmonella T3SS needle in 2012. Next, we did similar experimental analysis on the orthologues system in Shigella and we did homology modeling on other T3SS needles. Our studies corroborated that the T3SS needle which constitutes major part of the transport channel has very similar structure in different bacterial pathogens. Based on our results we started to mutate specific amino acids which could interact with substrate molecules during secretion. Using different assays we found that substrate secretion depends critically on the electrostatic and geometric properties of the inner surface of the needle. In collaboration with the lab of Helmut Grubmüller (MPI for Biophysical Chemistry) molecular dynamic simulations demonstrated the relevance of specific amino acids lining the channel surface for the transport process and explained how the unique channel architecture leads to efficient substrate secretion. Moreover, we showed, that substrate molecules might translocate in a right handed screw like pathway through the T3SS channel which is a remarkable feature of T3S. Our analysis provides for the first time insights into the substrate transport mechanism of T3SSs.

One of our aims was to characterize the cytoplasmic ring(c-ring) of the T3SS. We analyzed the assembly of a C-ring minimal core which is made by 4 different proteins. We solved the X-ray crystal structures of C-ring domains and we characterized how substrate molecules are recruited to this complex.

We also solved the X-ray crystal structures of proteins constituting the T3SS tip complex. To achieve this we used monoclonal antibody fragments in co-crystallization experiments. Structure of the tip protein domain may help to understand how host cells are manipulated by Shigella flexneri.

Reported by

Śledź nas na: RSS Facebook Twitter YouTube Zarządzany przez Urząd Publikacji UE W górę