[Display of the F18 fimbrial adhesin on the bacterial surface using type V secretion system and characterization of the binding domain of the adhesin].
The beta domains of autotransporters (also called type V secretion system) have been demonstrated to be feasible tools to display foreign passenger domain on the bacterial surface. In the present work, using the DNA manipulation the type V secretion system MisL displaying the binding doamin FedF1 of the F18ab fimbrial adhesion was constructed, and the full length adhesin FedF of F18ab fimbriae and the FedF mutant (with the 88th and 89th amino acid residues changed from Histadine to Alamine) on the surface of E. coli DH5alpha, respectively. The recombinant E. coli DH5alpha with the different recombinant plasmids of pnirBMisL-fedF1, pnirBMisL-fedF and pnirBMisL-fedF(M) were induced at anaerobic conditions. After induction, the binding doamin FedF1 of the FedF and adhesin FedF displayed on the surface of E. coli DH5alpha were tested for their agglutination and adhesion capability with the anti-rabbit sera against FedF subunit or the small intestinal epithelial cells from susceptible piglets. The results showed that the pnirBMisL-fedF1 or pnirBMisL-fedF recombinant bacteria could agglutinate with the anti-rabbit sera against FedF subunit and adhere to the small intestinal epithelial cells well. But the recombinant bacterial strain with the recombinant plasmid pnirBMisL-fedF(M) completely abolished the agglutination characteristic and receptor adhesiveness. These results confirmed that the binding doamin FedF1 of the F18ab fimbrial adhesin and the adhesin FedF of F18ab fimbriae could be transported and displayed functionally on the surface of E. coli DH5alpha by using type V secretion system and the His-88 and His-89 amino acid residues located in the FedF adhesin were important for the formation of the binding domain of the adhesin FedF of Fl8ab fimbriae.
Yuan
Zhi-wei
ZW
College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China.
Jiang
Ying
Y
Wang
Jian-ye
JY
Zhu
Guo-qiang
GQ
D018829:Adhesins, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000372:Agglutination Tests; D000818:Animals; D001422:Bacterial Adhesion; D001665:Binding Sites; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D016133:Polymerase Chain Reaction; D013552:Swine
D018829:Adhesins, Bacterial; D029968:Escherichia coli Proteins; C440653:FedF protein, E coli