Ribosome assembly studied by single-molecule force measurements
Thierry Bizebard (IBPC, Paris)
Ribosomes belong to the most complicated structures in biology. Their assembly is a question of fundamental interest, but is still poorly understood. In vitro reconstitution studies have shown that the ribosome assembly process is highly cooperative and starts with the binding of a few ribosomal (r-) proteins to rRNA, but how these early binders act is unknown. Our work focuses on the initial phase of the assembly of the large subunit (50S) of the E. coli ribosome, which involves 23S rRNA, five r-proteins and a selection of assembly “helper” proteins.
Our force measurements on single RNA molecules have allowed us to pinpoint several important properties of the early-binding r-proteins we have studied:
– These proteins bind with high cooperativity to the rRNA (as would be expected to obtain a high yield of fully assembled and active ribosomes).
– The r-proteins act as molecular clamps, stabilising the RNA 3D structure.
– As such, they afford a strong mechanical and energetical stabilisation of the ribonucleoprotein structure (which is also probably necessary for optimum activity).
In the near future, we intend to further improve the potential of our single-molecule measurements by implementing combined force/fluorescence manipulations, and apply this methodology to our study of the early phase of E. coli large ribosomal subunit assembly.