A 20-kilodalton N-terminal fragment of the D15 protein contains a protective epitope(s) against Haemophilus influenzae type a and type b.
A conserved 80-kDa minor outer membrane protein, D15, of Haemophilus influenzae has been shown to be a protective antigen in laboratory animals against H. influenzae type a (Hia) or type b (Hib) infection. To localize the protective B-cell epitope(s) within the D15 protein and to further explore the possibility of using synthetic peptides as vaccine antigens, a 20-kDa N-terminal fragment of D15 protein (truncated D15 [tD15]) was expressed as a fusion protein with glutathione S-transferase in Escherichia coli. The tD15 moiety was cleaved from glutathione S-transferase by using thrombin and purified to homogeneity. The purified soluble tD15 appeared to contain immunodominant protective epitope(s) against Hia and Hib, since rabbit antisera directed against tD15 were capable of protecting infant rats from Hia or Hib bacteremia. The ease of purification of soluble tD15, therefore, makes it a better candidate antigen than the full-length recombinant D15 which is produced as inclusion bodies in E. coli. Furthermore, both the purified tD15 fragment and a mixture of tD15-derived peptides spanning amino acid residues 93 to 209 of the mature D15 protein were capable of inhibiting the protection against Hib conferred on infant rats by rabbit anti-tD15 antiserum, indicating that the protective epitopes of D15 may not be conformational. However, the administration of pooled rabbit immune sera raised against the same panel of peptides failed to protect infant rats from Hib infection.
Yang
Y p
Yp
Research Center, Pasteur Merieux Connaught Canada, North York, Ontario, Canada M2R 3T4. ypyang@ca.pmc-vacc.com
Thomas
W R
WR
Chong
P
P
Loosmore
S M
SM
Klein
M H
MH
D000595:Amino Acid Sequence; D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000276:immunology* / Q000302:isolation & purification; D018985:Epitopes, B-Lymphocyte; D006193:Haemophilus influenzae / Q000737:chemistry / Q000276:immunology*; D007106:Immune Sera / Q000276:immunology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000276:immunology* / Q000302:isolation & purification; D011817:Rabbits; D051381:Rats; D017208:Rats, Wistar; D014614:Vaccines, Synthetic / Q000276:immunology
D001425:Bacterial Outer Membrane Proteins; D018985:Epitopes, B-Lymphocyte; D007106:Immune Sera; D010446:Peptide Fragments; D014614:Vaccines, Synthetic