Expression of thePlasmodium falciparumImmunodominant Epitope (NANP)₄on the Surface ofSalmonella entericaUsing the Autotransporter MisL

Abstract

Gram-negative bacterial proteins which are exported from the cytosol to the external environment by the type V secretion system are also known as autotransporters. Once translocated to the periplasmic compartment by the sec-dependent general secretory pathway, their C-terminal domain forms a pore through which the N-terminal domain travels to the outer membrane without the need of other accessory proteins. MisL (protein of membrane insertion and secretion) is a protein of unknown function located in the pathogenicity island SPI-3 of Salmonella enterica and classified as an autotransporter due to its high homology to Escherichia coli AIDA-I. In the present work, the MisL C-terminal translocator domain was used to display the immunodominant B-cell epitope of the circumsporozoite protein (CSP) from Plasmodium falciparum on the surface of Salmonella enterica serovar Typhimurium (serovar Typhimurium SL3261) and serovar Typhi (serovar Typhi CVD 908). The MisL ␤ domain was predicted by alignment with AIDA-I, amplified from serovar Typhimurium SL3261, cloned in a plasmid fused to four repeats of the tetrapeptide NANP behind the Escherichia coli heat-labile enterotoxin B subunit signal peptide to ensure periplasmic traffic, and expressed under the control of the anaerobically inducible nirB promoter. The fusion protein was translocated to the outer membrane of both bacterial strains, although the foreign epitope was displayed more efficiently in serovar Typhimurium SL3261, which elicited a better specific antibody response in BALB/c mice. More importantly, antibodies were able to recognize the native CSP in P. falciparum sporozoites. These results confirm that MisL is indeed an autotransporter and that it can be used to express foreign immunogenic epitopes on the surface of gramnegative bacteria.

Autodisplay

“…Recently, use of E. coli outer membrane protein TolC for surface display of protective listerial B-and T-cell epitopes in a Salmonella vaccine strain has been shown to be efficacious in vivo (46). In summary, autotransporters facilitate effective surface display of antigenic determinants on live Salmonella vaccine carriers, leading to humoral (42) and cellular immune responses in vivo (29). This study demonstrated that surface display of a Helicobacter antigen via the AIDA-I autotransporter can induce protective immune responses.…”

“…Humoral immune responses have been observed after vaccination of mice with attenuated Salmonella strains with immunogenic determinants exposed on their surfaces by means of outer membrane proteins from E. coli, like OmpA (17), PhoE (24,47), LamB (6,18,32), and P87 fimbriae (7), the ice-nucleating protein from Pseudomonas aeruginosa (33), the main flagellar component FliC from Salmonella (9,40,40), or the Salmonella autotransporter MisL (42). Recently, use of E. coli outer membrane protein TolC for surface display of protective listerial B-and T-cell epitopes in a Salmonella vaccine strain has been shown to be efficacious in vivo (46).…”

Salmonella enterica Serovar Typhi Ty21a Expressing Human Papillomavirus Type 16 L1 as a Potential Live Vaccine against Cervical Cancer and Typhoid Fever

“…To our knowledge, experiments with recombinant Ty800 in the nasal murine model were not previously reported, while plasmid stability was observed in both Ty21a (16) and CVD908-htrA (27). The instability of L1S plasmids in CVD908-htrA may be linked to the presence of a constitutive promoter in our plasmids instead of the in vivo-inducible nirB promoter, which was generally used in other recombinant CVD908-htrA strains (27,44,50).…”

MarT Activates Expression of the MisL Autotransporter Protein of Salmonella enterica Serotype Typhimurium

“…The MisL C-terminal translocator domain can be used to display foreign antigens and elicits specific antibody responses in mice (30,31,36). Mice infected with serotype Typhimurium seroconvert to MisL, which provides indirect evidence for in vivo expression of this autotransporter protein (5).…”