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The minimal Tat translocase

(2009) Ploeg, René van der

Thesis UNDER EMBARGO! Till november 2011

Bacteria use specific transport system for uptake and secretion of molecules with functions in metabolism, defence against other organisms and communication between cells. The purpose of some transport systems is the secretion of proteins. Research on protein secretion in Grampositive bacteria is commonly performed with the model organism Bacillus subtilis. The high protein secretion capacity of this bacterium makes it applicable in the biotechnological industry as a “cell factory” for valuable secreted proteins. This accounts mostly for proteins from bacilli. Secretion of proteins from other organisms is often difficult. More research is needed to increase the applicability of B. subtilis. The present studies focus on a system that can transport folded proteins. The transported proteins carry a molecular zip-code, the signal peptide, that harbours two neighbouring arginine residues. Therefore the system is named the “Twin arginine translocase”, or in short Tat. The TatAC system in B. subtilis represents a minimal variant of this system, because only two components are necessary for activity. The results show that it is possible to target proteins for secretion via the B. subtilis Tat system. Novel insights are that internal and external factors can influence the use of Tat. High-level overproduction of the esterase LipA results in its secretion via Tat. Also, the salinity of the growth medium is important for Tat-dependent secretion. This is apparently to be due to an intrinsic salt-sensitivity of the Tat system. Overall, this PhD research provides insights in the function and application potential of the B. subtilis Tat system.