Ribosome-catalyzed synthesis of proteins according to the genetic information stored as DNA is fundamental to all living cells. Proteins perform practically all the daily work of a cell, such as transporting molecules from one part of the cell to another and, particularly, catalyzing and regulating all chemical reactions taking place in the cell. The molecular machinery of protein synthesis has been characterized in great detail, mainly using reconstituted systems where the individual components have been purified from cells and then studied in the test tube. However, whereas powerful to dig out the details, this reductionist approach can never account for all the aspects of the complex, crowded environment of the living cell. The aim of this project is to fit the detailed pieces of information on the process of protein synthesis into a bigger picture taking into consideration the full plethora of players present in the cell. In particular, we want to understand how the sequence of the template, the mRNA, determines how, when, and where the different proteins should be folded, and, in particular, how these decisions are made during ongoing protein synthesis. This information will be particularly important for optimization of protein production, such as cost-effective production of therapeutic proteins using bacterial cells. To achieve this, we are developing fluorescence-based tools to study individual molecules, one by one, directly during action, inside living E. coli cells.
The project is divided into three main subprojects:
• Subproject 1: How are mRNA translation rates tuned, globally and locally, to allow for rapid polypeptide production, whilst still maintaining proper polypeptide folding and/or targeting to non-cytosolic compartments?
• Subproject 2: What is the timing and capacity of Signal Recognition Particle (SRP)-mediated cotranslational targeting of nascent polypeptides to the membrane-bound peptide translocation complexes (i.e. the translocons)?
• Subproject 3: How are mRNA translation rates affected, globally and locally, by ribosome-targeting antibiotic drugs, and how is this connected to the drug’s bacteriostatic/bactericidal effect?
