Ribosome Assembly in Bacteria
A major focus our laboratory is the dynamics of assembly the ribosome, which is the large macromolecular machine responsible for protein synthesis in all cells. The ribosome is ~2.7 MDa, and is composed of three large ribosomal RNA molecules and 55 ribosomal proteins. Ribosome assembly in bacteria occurs co-transcriptionally at highly specialized loci that are sites of this major metabolic activity that accounts for about 1/3 of the energy budget of a rapidly dividing cell.
Key questions about ribosome assembly focus on the structure of assembly intermediates, the role of > 30 asssembly cofactors, the sequence of RNA conformational changes that occurs, and the coordination of binding of the ribosomal proteins. Ribosome assembly is incredibly efficient, requiring only ~2 minutes, which is only somewhat longer than the transcription time for the ribosomal RNA.
We use quantitative mass spectrometry to determine the protein composition of assembly intermediates in conjunction with electron microscopy to determine the structure of those intermediates. In addition, we use fluorescence methods to monitor protein binding and RNA conformational changes during assembly.
Overall, we are working to define the series of events that will define the ribosome assembly pathway in terms of a molecular movie of transcription, RNA folding, RNA processing, protein binding and cofactor assistance that ultimately produces the mature ribosome.