Translocation of jellyfish green fluorescent protein via the Tat system ofEscherichia coli and change of its periplasmic localization in response toosmotic up-shock

The bacterial twin arginine translocation (Tat) pathway is capable of expor ting cofactor-containing enzymes into the periplasm. To assess the capacity of the Tat pathway to export heterologous proteins and to gain information about the property of the periplasm, we fused the twin arginine signal pep tide of the trimethyl-amine N-oxide reductase to the jellyfish green fluore scent protein (GFP), Unlike the Sec pathway, the Tat system successfully ex ported correctly folded GFP into the periplasm of Escherichia coli. Interes tingly, GFP appeared as a halo in most cells and occasionally showed a pola r localization in wild type strains. When subjected to a mild osmotic up-sh ock, GFP relocalized very quickly at the two poles of the cells. The conver sion from the halo structure to a periplasmic gathering at particular locat ions was also observed with spherical cells of the Delta rodA-pbpA mutant o r of the wild type strain treated with lysozyme. Therefore, the periplasm i s not a uniform compartment and the polarization of GFP is unlikely to be c aused by simple invagination of the cytoplasmic membrane at the poles. More over, the polar gathering of GFP is reversible; the reversion was accelerat ed by glucose and inhibited by azide and carbonyl cyanide m-chlorophenylhyd razone, indicating an active adaptation of the bacteria to the osmolarity i n the medium. These results strongly suggest a relocalization of periplasmi c substances in response to environmental changes. The polar area might be the preferential zone where bacteria sense the change in the environment.