Molecular cloning, sequencing, expression, and characterization of an immunogenic 43-kilodalton lipoprotein of Bartonella bacilliformis that has homology to NlpD/LppB.
A recombinant clone expressing an immunoreactive antigen of Bartonella bacilliformis was isolated by screening a genomic DNA library with serum from a patient with the chronic verruga phase of bartonellosis. The clone, pBIPIM-17, contained a partial open reading frame that expressed an immunoreactive fusion protein. Subsequent rescreening of the library by plaque hybridization resulted in the isolation of recombinant clones that contain the entire open reading frame. The open reading frame (ORF-401) is capable of encoding a protein of 401 amino acids with a predicted molecular mass of 43 kDa. The deduced amino acid sequence of the encoded protein was found to be highly homologous to a recently identified bacterial lipoprotein (LppB/NlpD) which has been associated with virulence. Evidence has been provided to show that the 43-kDa antigen of B. bacilliformis is a lipoprotein and that it is likely to use the same biosynthetic pathway as other bacterial lipoproteins. This is the first report to date that characterizes a lipoprotein of B. bacilliformis. The immunogenicity of the B. bacilliformis LppB homologue was demonstrated by Western blot analysis using sera from patients with clinical bartonellosis. Sera from patients who had a high titer for Bartonella henselae, the causative agent of bacillary angiomatosis and cat scratch disease, also recognized the recombinant 43-kDa antigen, suggesting that a homologue of this antigen is present in B. henselae. Using a cocktail of synthetic peptides corresponding to predicted major antigenic sites, polyclonal antiserum specific for the LppB homologue of B. bacilliformis was generated. This antiserum did not recognize the NlpD homologue of Escherichia coli or the 43-kDa antigen of B. henselae.
Padmalayam
I
I
Division of Viral and Rickettsial Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30333, USA. ixp0@cdc.gov
Kelly
T
T
Baumstark
B
B
Massung
R
R
D000595:Amino Acid Sequence; D000818:Animals; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology*; D001426:Bacterial Proteins; D001473:Bartonella / Q000276:immunology*; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D029968:Escherichia coli Proteins; D007106:Immune Sera / Q000276:immunology; D008074:Lipoproteins / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011499:Protein Processing, Post-Translational; D011817:Rabbits; D017386:Sequence Homology, Amino Acid
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D007106:Immune Sera; D008074:Lipoproteins; C490122:NlpD protein, E coli; C086389:NlpD protein, bacteria