title: Effect of lipid modification on the physicochemical, structural, antigenic and immunoprotective properties of Haemophilus influenzae outer membrane protein P6.
abstract: The outer membrane lipoprotein, P6 of Haemophilus influenzae was studied to determine the importance of the native palmitoyl moiety on its physicochemical and immunological properties. A recombinant P6 (rP6) molecule devoid of lipidation signal sequence was expressed in Escherichia coli and its properties were compared to those of the palmitylated protein purified from H. influenzae. The isoelectric point of rP6 was more acidic than that of the native protein and also exhibited less secondary structure than P6 as judged by circular dichroism. However, both forms of P6 induced identical P6-specific antibody titers in guinea pigs when Freund's adjuvant was used. These antisera reacted with a panel of overlapping P6 peptides in a comparable manner and in addition, rabbit antisera raised against the P6 peptides reacted equally well with P6 and rP6. Furthermore, all human convalescent sera tested exhibited similar anti-P6 and anti-rP6 antibody titers. However, rP6 was less immunogenic than P6 when administered either without adjuvant or in alum and when tested in competitive inhibition studies with anti-P6 antibodies, was a less effective inhibitor than native P6, suggesting a diminution in some of the antigenic activity of rP6. In spite of these differences, rP6 was capable of eliciting a protective antibody response against live H. influenzae type b challenge in a modified infant rat model of bacteremia. These findings demonstrate that the non-fatty acylated rP6 could possibily be substituted for native P6 in a vaccine against H. influenzae.
authors.affiliation: Research Center, Pasteur Merieux, Connaught, Ontario, Canada.
authors.forename: Y P
authors.initials: YP
authors.lastname: Yang
authors.forename: R S
authors.initials: RS
authors.lastname: Munson
authors.forename: S
authors.initials: S
authors.lastname: Grass
authors.forename: P
authors.initials: P
authors.lastname: Chong
authors.forename: R E
authors.initials: RE
authors.lastname: Harkness
authors.forename: L
authors.initials: L
authors.lastname: Gisonni
authors.forename: O
authors.initials: O
authors.lastname: James
authors.forename: Y
authors.initials: Y
authors.lastname: Kwok
authors.forename: M H
authors.initials: MH
authors.lastname: Klein
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology; D000918:Antibody Specificity / Q000276:immunology; D000942:Antigens, Bacterial / Q000276:immunology*; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000276:immunology* / Q000627:therapeutic use; D001431:Bacteriological Techniques; D001667:Binding, Competitive / Q000276:immunology; D003602:Cytotoxicity, Immunologic / Q000276:immunology; D005260:Female; D006168:Guinea Pigs; D006192:Haemophilus Infections / Q000276:immunology / Q000517:prevention & control*; D018073:Haemophilus Vaccines / Q000737:chemistry* / Q000276:immunology* / Q000627:therapeutic use; D006193:Haemophilus influenzae / Q000276:immunology*; D006801:Humans; D008055:Lipids / Q000737:chemistry / Q000276:immunology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011817:Rabbits; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification / Q000627:therapeutic use
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D018073:Haemophilus Vaccines; D008055:Lipids; C069990:P6 outer membrane protein, Haemophilus; D011994:Recombinant Proteins