C-TERMINAL GLYCINE-HISTIDINE TAGGING OF THE OUTER-MEMBRANE PROTEIN IGA-BETA OF NEISSERIA-GONORRHOEAE

A glycine-histidine tag (Gly,His,) was added to the C-terminus of a fu sion protein consisting of the cholera toxin B-subunit (CtxB) and the IgA protease beta-domain (Iga beta). The aim was to facilitate single- step purification and to create a suitable tool for kinetic and struct ural studies on Iga beta-driven protein translocation across the outer membrane of Gram-negative bacteria. We demonstrate that the glycine-h istidine tag does not interfere with the assembly of Iga beta in the o uter membrane and that the translocator function of the modified Iga b eta is maintained. The applicability of the new construct for the diss ection of the Iga beta mediated translocation process and general aspe cts of C-terminal histidine tagging of outer membrane proteins are dis cussed.