Expression of heterologous peptides at two permissive sites of the MalE protein: antigenicity and immunogenicity of foreign B-cell and T-cell epitopes.
We previously determined a number of 'permissive' sites in the periplasmic maltose-binding protein (MalE) from Escherichia coli. These sites accept the insertion of heterologous peptides without major deleterious consequences for the activities, structure and cellular location of the protein. This study explores the versatility of two such permissive sites for the synthesis of foreign peptides, and examines the antigenicity and the immunogenicity of the inserts. One site is located after amino acid 133 (aa133) of MalE, and the other after aa303. Both sites tolerate inserts of up to at least 70 aa and accept sequences of different natures. Hydrophobic aa sequences are accepted, although strongly hydrophobic sequences, such as the Sendai virus F protein membrane anchor, affected export. We compared the antigenic and the immunogenic properties of peptides derived from the coat proteins of HBV and poliovirus which contain well defined B-cell epitopes. Specific monoclonal antibodies show that the antigenic properties of the inserted B-cell epitopes were different at the two sites. Despite these differences, the inserted peptides elicited strong and comparable antibody responses in mice against the corresponding synthetic peptides. In this case, and with these criteria, the molecular context of the peptides did not affect the immunogenicity of B-cell epitopes. We show for the first time that when a foreign peptide carrying a T-cell epitope was inserted in MalE, the hybrid proteins can elicit a T-cell response against the foreign peptide both in vivo and in vitro. Furthermore, the MalE hybrid was as efficient as free peptide in stimulating T-cell hybridomas in vitro. The MalE vectors provide a powerful genetic system to study how the position and the conformation of a peptide within a protein affect the B-cell and T-cell responses.
Martineau
P
P
U.P.M.T.G. (CNRS UA1444), Paris, France.
Guillet
J G
JG
Leclerc
C
C
Hofnung
M
M
D018528:ATP-Binding Cassette Transporters; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D001402:B-Lymphocytes / Q000276:immunology*; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D006514:Hepatitis B Surface Antigens / Q000276:immunology; D008320:Maltose / Q000378:metabolism; D058619:Maltose-Binding Proteins; D008961:Models, Structural; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D033902:Periplasmic Binding Proteins; D010957:Plasmids; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000276:immunology; D015183:Restriction Mapping; D013601:T-Lymphocytes / Q000276:immunology*
D018528:ATP-Binding Cassette Transporters; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D002352:Carrier Proteins; D000939:Epitopes; D029968:Escherichia coli Proteins; D006514:Hepatitis B Surface Antigens; C461619:MalE protein, E coli; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D033902:Periplasmic Binding Proteins; D011994:Recombinant Proteins; C461599:maltose transport system, E coli; D008320:Maltose