The twin-arginine signal peptide of PhoD and the TatAd/Cd proteins of Bacillus subtilis form an autonomous Tat translocation system.
The bacterial twin-arginine translocation (Tat) pathway has been recently described for PhoD of Bacillus subtilis, a phosphodiesterase containing a twin-arginine signal peptide. The expression of phoD is co-regulated with the expression of tatA(d) and tatC(d) genes localized downstream of phoD. To characterize the specificity of PhoD transport further, translocation of PhoD was investigated in Escherichia coli. By using gene fusions, we analyzed the particular role of the signal peptide and the mature region of PhoD in canalizing the transport route. A hybrid protein consisting of the signal peptide of beta-lactamase and mature PhoD was transported in a Sec-dependent manner indicating that the mature part of PhoD does not contain information canalizing the selected translocation route. Pre-PhoD, as well as a fusion protein consisting of the signal peptide of PhoD (SP(PhoD)) and beta-galactosidase (LacZ), remained cytosolic in the E. coli. Thus, SP(PhoD) is not recognized by E. coli transport systems. Co-expression of B. subtilis tatA(d)/C(d) genes resulted in the processing of SP(PhoD)-LacZ and periplasmic localization of LacZ illustrating a close substrate specificity of the TatA(d)/C(d) transport system. While blockage of the Sec-dependent transport did not affect the localization of SP(PhoD)-LacZ, translocation and processing was dependent on the pH gradient of the cytosolic membrane. Thus, the minimal requirement of a functional Tat-dependent protein translocation system consists of a twin-arginine signal peptide-containing Tat substrate, its specific TatA/C proteins, and the pH gradient across the cytosolic membrane.
Pop
Ovidiu
O
Institutes of Molecular Biology, Jena University, Winzerlaer Str. 10, D-07745 Jena, Germany.
Martin
Ulrike
U
Abel
Christian
C
Müller
Jörg P
JP
D001120:Arginine; D001412:Bacillus subtilis / Q000235:genetics*; D001692:Biological Transport / Q000235:genetics; D002876:Chromosomes, Bacterial / Q000235:genetics; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins / Q000235:genetics*; D010957:Plasmids; D021382:Protein Sorting Signals / Q000235:genetics*; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity; D014178:Translocation, Genetic; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D017931:DNA Primers; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D021382:Protein Sorting Signals; C408585:twin-arginine translocase complex, E coli; D001120:Arginine; D001616:beta-Galactosidase