C-terminal glycine-histidine tagging of the outer membrane protein Iga beta of Neisseria gonorrhoeae.
A glycine-histidine tag (Gly3His6) was added to the C-terminus of a fusion 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 structural studies on Iga beta-driven protein translocation across the outer membrane of Gram-negative bacteria. We demonstrate that the glycine-histidine tag does not interfere with the assembly of Iga beta in the outer membrane and that the translocator function of the modified Iga beta is maintained. The applicability of the new construct for the dissection of the Iga beta mediated translocation process and general aspects of C-terminal histidine tagging of outer membrane proteins are discussed.
Strauss
A
A
Max-Planck-Institut für Biologie, Abteilung Infektionsbiologie, Tübingen, Germany.
Pohlner
J
J
Klauser
T
T
Meyer
T F
TF
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics*; D001483:Base Sequence; D001693:Biological Transport, Active; D002462:Cell Membrane / Q000378:metabolism; D002772:Cholera Toxin / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics; D004151:Dipeptides / Q000235:genetics; D008969:Molecular Sequence Data; D009344:Neisseria gonorrhoeae / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics; D016324:Sequence Tagged Sites; D012697:Serine Endopeptidases / Q000235:genetics*
D001425:Bacterial Outer Membrane Proteins; D004269:DNA, Bacterial; D004151:Dipeptides; D011993:Recombinant Fusion Proteins; C027787:glycylhistidine; D002772:Cholera Toxin; D012697:Serine Endopeptidases; C021433:IgA-specific serine endopeptidase