Antigen 43-mediated autotransporter display, a versatile bacterial cell surface presentation system.
Antigen 43 (Ag43), a self-recognizing outer membrane protein of Escherichia coli, has been converted into an efficient and versatile tool for surface display of foreign protein segments. Ag43 is an autotransporter protein characterized by the feature that all information required for transport to the outer membrane and secretion through the cell envelope is contained within the protein itself. Ag43 consists of two subunits (alpha and beta), where the beta-subunit forms an integral outer membrane translocator to which the alpha-subunit is noncovalently attached. The simplicity of the Ag43 system makes it ideally suited as a surface display scaffold. Here we demonstrate that the Ag43 alpha-module can accommodate and display correctly folded inserts and has the ability to display entire functional protein domains, exemplified by the FimH lectin domain. The presence of heterologous cysteine bridges does not interfere with surface display, and Ag43 chimeras are correctly processed into alpha- and beta-modules, offering optional and easy release of the chimeric alpha-subunits. Furthermore, Ag43 can be displayed in many gram-negative bacteria. This feature is exploited for display of our chimeras in an attenuated Salmonella strain.
Kjaergaard
Kristian
K
Microbial Adhesion Group, Section of Molecular Microbiology, BioCentrum-DTU, Technical University of Denmark, DK-2800 Lyngby, Denmark.
Hasman
Henrik
H
Schembri
Mark A
MA
Klemm
Per
P
D018829:Adhesins, Bacterial / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D018830:Adhesins, Escherichia coli; D000942:Antigens, Bacterial / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015153:Blotting, Western; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D029968:Escherichia coli Proteins; D036781:Fimbriae Proteins; D005455:Fluorescent Antibody Technique; D037102:Lectins; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011995:Recombination, Genetic; D019779:Salmonella enterica / Q000235:genetics
D018829:Adhesins, Bacterial; D018830:Adhesins, Escherichia coli; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D000939:Epitopes; D029968:Escherichia coli Proteins; D037102:Lectins; D011993:Recombinant Fusion Proteins; C118376:antigen 43, E coli; C058769:fimH protein, E coli; D036781:Fimbriae Proteins