Characterisation of the twin-arginine protein translocation pathway of Bacillus subtilis
It has been demonstrated previously that phosphodiesterase PhoD containing a double arginine signal peptide, is transported Tat dependent. The studies presented in the thesis demonstrate that PhoD and the transport components TatAd and TatCd form an autonomous protein translocation system in the donor strain and in heterologous organisms. It mediates the transport of PhoD a s well as hybrid proteins containing the signal peptide of PhoD as translocation signal. Particularly, it was shown that the twin-arginine signal peptide of PhoD canalised the Tat-dependent transport of beta-galactosidase LacZ in E. coli and the E. coli phytase AppA in B. subtilis. Biochemical investigations demonstrated that a protein with one calculated membrane spanning domain is localised in the membrane as well as soluble in the cytosol. The soluble forms homo-multimeric particles which have affinity to prePhoD, particularly to its double arginine motif of the signal peptide. Data suggest that TatAd could play a role as targeting factor for PhoD. In addition, was found to form multimeric complexes in the membranes of reconstituted liposomes with affinity for prePhoD. Therefore it can be concluded that TatAd may significantly contribute to the formation of the translocation channel, which conducts PhoD through the cytosolic membrane.