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ERC

assemblyNMR Report Summary

Project ID: 337490
Funded under: FP7-IDEAS-ERC
Country: Germany

Mid-Term Report Summary - ASSEMBLYNMR (3D structures of bacterial supramolecular assemblies by solid-state NMR)

In this project we focused on the characterization of protein structure, dynamics and function of three different bacterial systems of interest: The bacteria-specific cytoskeletal element bactofilin (Caulobacter crescentus: subunit name BacA), the type i pilus rod (Uropathogenic E. coli: subunit name FimA), and the type iii secretion system needle (Shigella flexneri: subunit name MxiH). Those systems have in common that the respective subunits polymerize and form supramolecular assemblies that may be part of complex molecular machines. As a consequence of their size and insolubility they pose a significant challenge to the standard techniques for 3D protein structure determination at atomic resolution, which are X-ray crystallography and solution NMR. In contrast, solid-state NMR neither requires solubility nor long-range order and can thus in principle be readily applied to those systems. It was a major goal of this project to demonstrate that structure, dynamics and function of such systems can indeed be characterized by solid-state NMR, possibly in combination with data from cryo-electron microscopy (cryo-EM). Cryo-EM has recently witnessed tremendous progress and it perfectly complements solid-state NMR data (which provide local information on secondary structure, subunit tertiary structure and subunit-subunit intermolecular restraints) by providing global information on the assembly (rigid body positioning of the subunits, symmetry, diameter etc.).

In the first funding period we determined the structures of bactofilin and of the type i pilus rod and thereby demonstrated the power of solid-state NMR for the study of supramolecular protein assemblies. In both cases we complemented the data from solid-state NMR with data from other structural methods (solution NMR, electron microscopy). It is now increasingly recognized that hybrid approaches are important tools to address the structures of complex molecular machines and with our studies we could increase the knowledge in this growing research field.

The structure of the type i pilus yielded valuable information on the atomic scale, but in general corroborated/refined existing earlier coarse models. In contrast, the bactofilin structure revealed a very surprising beta-helical fold with six windings. Such a beta-helical architecture had never been observed before for other cytoskeletal filaments. Interestingly, however, the structure bears similarities to that of the fungal prion protein HET-s.

Bactofilins are a new class of cytoskeletal proteins that are involved in key cellular processes. For instance, in the human pathogen Helicobacter pylori, they are responsible for maintaining its characteristic helical cell shape, a feature required for cells to efficiently colonize the gastric mucus. Bactofilins only occur in bacteria but not in humans and thus may be an interesting pharmacological target. The here determined structure could be helpful for such endeavors.

Contact

Anne Höner, (EU Liaison Officer)
Tel.: +49 30 94793286
Fax: +49 30 94793109
E-mail
Record Number: 194473 / Last updated on: 2017-02-15
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