Biological activity of serum antibodies to a nonacylated form of lipoprotein D of Haemophilus influenzae.
Protein D, a surface-exposed 42-kDa membrane lipoprotein, is well conserved among both type b and nontypeable Haemophilus influenzae strains, and it is considered a vaccine against H. influenzae infections. Here, we report the large-scale purification of a nonacylated form of protein D (PDm) from the periplasmic space of Escherichia coli overexpressing PDm. Screening of human sera for levels of antibodies to PDm demonstrated that the immunoglobulin G (IgG) antibody level is above background levels in infants less than 6 months of age. Following a drop to background values in the age group 6 months to 1 year, IgG antibody levels start to increase, together with IgA antibody levels, after 1 year of age. The first appearance of serum IgM antibodies is in 6-month- to 1-year-old infants whose IgG antibody levels have dropped to the postnatal background level. Affinity-purified antibodies from humans and from PDm-immunized rats detected epitopes of protein D which are normally exposed on the bacterial surface. Affinity-isolated human anti-PDm antibodies eluted in acidic buffer were not bactericidal against H. influenzae. Loss of bactericidal activity may occur in this buffer, as was demonstrated in pooled human sera with high bactericidal activity after incubation in the same buffer. Hyperimmunization of rats with PDm induced high levels of serum IgG and IgA antibodies against PDm and significant bactericidal activity against homologous and heterologous H. influenzae strains.
Department of Medical Microbiology, Lund University, Malmö University Hospital, Sweden. Mustafa.Akkoyunlu@mikrobiol.mas.lu.se.
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Janson
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Ruan
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Forsgren
D000215:Acylation; D000293:Adolescent; D000368:Aged; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood / Q000276:immunology* / Q000302:isolation & purification; D001426:Bacterial Proteins; D002352:Carrier Proteins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D002648:Child; D002675:Child, Preschool; D002846:Chromatography, Affinity; D006193:Haemophilus influenzae / Q000276:immunology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007114:Immunization; D007070:Immunoglobulin A / Q000097:blood / Q000302:isolation & purification; D007072:Immunoglobulin D; D007074:Immunoglobulin G / Q000097:blood / Q000302:isolation & purification; D007075:Immunoglobulin M / Q000097:blood / Q000302:isolation & purification; D007223:Infant; D008074:Lipoproteins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D008875:Middle Aged; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology / Q000302:isolation & purification
D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D002352:Carrier Proteins; D007070:Immunoglobulin A; D007072:Immunoglobulin D; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D008074:Lipoproteins; D011994:Recombinant Proteins; C067989:glpQ protein, Haemophilus influenzae