Monoclonal antibodies with specificities for Streptococcus pneumoniae group 9 capsular polysaccharides

Kolberg, Jan

doi:10.1016/s0928-8244(98)00018-2

Cited by 5 publications

(5 citation statements)

References 13 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Measurements were performed as described previously (Kolberg & Jones, 1998). Briefly, flat-bottomed microtitre plates (Nunc Immunoplate) were coated overnight at room temperature with recombinant enolase protein (2 mg ml 21 ) in PBS.…”

Section: Methodsmentioning

confidence: 99%

Streptococcus pneumoniae enolase is important for plasminogen binding despite low abundance of enolase protein on the bacterial cell surface

et al. 2006

View full text Add to dashboard Cite

Enolase represents one of the anchorless surface proteins of Streptococcus pneumoniae and has previously been identified as a plasminogen-binding protein, endowing this pathogen with host proteolytic activity. In this study the mAb 245,C-6 (IgG1) was produced in a BALB/c mouse after immunizing with a protein fraction from S. pneumoniae. The mAb reacted with recombinant pneumococcal enolase both under non-denaturing and denaturing conditions. The epitope for the mAb was mapped to residues 55 DKSRYGGLG 63 of pneumococcal enolase using a peptide array. By applying the previously reported structure of enolase, this epitope was localized in a surface-exposed loop in each of the monomers of the octameric enolase. Previous immunoelectron microscopic studies, using polyclonal rabbit antibodies against enolase, depicted enolase on the cell surface but did not quantify the amount of surface-exposed enolase on viable pneumococci. Here, flow cytometry revealed no binding of mAb 245,C-6 to viable pneumococci, including TIGR4 and its non-encapsulated isogenic mutant, and only a minor increase of fluorescence intensity was measured when the polyclonal anti-enolase antibodies were used. In contrast, control antibodies recognizing the choline-binding proteins (CBPs) PspA and PspC showed high reactivities. The non-encapsulated TIGR4 did not show increased levels of antibody binding for mAb 245,C-6 or polyclonal anti-enolase antibodies, but revealed increased binding of polyclonal antibodies reacting with PspA or PspC. These results suggest that, compared to other surface-exposed proteins such as CBPs, the amount of enolase under the selected conditions is low. Flow cytometry, however, with FITC-labelled plasminogen demonstrated that the amount of surface-exposed enolase is important for plasminogen binding and, therefore, is also important for pneumococcal pathogenesis. INTRODUCTIONStreptococcus pneumoniae is an important human pathogen causing a variety of diseases including pneumonia, meningitis, sepsis and otitis media. A prerequisite for invasive pneumococcal diseases (infections) is the ability of the bacteria to colonize the mucosal surface and penetrate the mucosal barriers. During these processes binding of bacterial factors such as adhesins are essential for the interactions with receptors on host cells. Among the bacterial factors that mediate direct adherence to human cells are phosphorylcholine (Cundell et al., 1995) and the choline-binding protein PspC, also known as SpsA and CbpA (Brooks-Walter et al., 1999;Hammerschmidt et al., 1997;Rosenow et al., 1997). The PspC protein binds to the secretory component of the polymeric Ig receptor or secretory IgA and this interaction mediates transmigration of pneumococci through mucosal epithelial cells (Elm et al., 2004;Zhang et al., 2000). In addition, PspC and the PspClike protein Hic have been shown to interact with complement factor H (Dave et al., 2004;Janulczyk et al., 2000). Many pathogenic bacteria also produce receptors for Abbreviations: GAPDH, glyceraldehyde-3-phos...

show abstract

Section: Methodsmentioning

confidence: 99%

Streptococcus pneumoniae enolase is important for plasminogen binding despite low abundance of enolase protein on the bacterial cell surface

et al. 2006

View full text Add to dashboard Cite

show abstract

“…The measurements were performed as previously described [23]. Briefly, flat‐bottomed microtiter plates (Nunc‐Immunoplate, Nunc, Denmark) were coated overnight at room temperature with recombinant proteins 1 μg ml −1 in phosphate‐buffered saline.…”

Section: Methodsmentioning

confidence: 99%

Epitope mapping of pneumococcal surface protein A of strain Rx1 using monoclonal antibodies and molecular structure modelling

Kolberg¹,

Aase²,

Rødal³

et al. 2003

FEMS Immunology &Amp; Medical Microbiology

View full text Add to dashboard Cite

Pneumococcal surface protein A (PspA) is an antigenic variable vaccine candidate of Streptococcus pneumoniae. Epitope similarities between PspA from the American vaccine candidate strain Rx1 and Norwegian clinical isolates were studied using PspA specific monoclonal antibodies (mAbs) made against clinical Norwegian strains. Using recombinant PspA/Rx1 fragments and immunoblotting the epitopes for mAbs were mapped to two regions of amino acids, 1-67 and 67-236. The discovered epitopes were visualized by modelling of the PspA:Fab part of mAb in three dimensions. Flow cytometric analysis showed that the epitopes for majority of mAbs were accessible for antibody binding on live pneumococci. Also, the epitopes for majority of the mAbs are widely expressed among clinical Norwegian isolates.

show abstract

“…Further fractionation of the soluble proteins were done by preparative SDS–PAGE. Proteins in the molecular mass region of 46–70 kDa were electroeluted, dialysed against PBS and used for immunisation of BALB/c mice, essentially as previously described [14]. Spleen cells were fused with NSO myeloma cells by standard methods.…”

Section: Methodsmentioning

confidence: 99%

Streptococcus pneumoniaeheat shock protein 70 does not induce human antibody responses during infection

Kolberg

Høiby²,

Aase

et al. 2000

FEMS Immunology &Amp; Medical Microbiology

View full text Add to dashboard Cite

Mouse monoclonal antibodies (mAbs) were developed against Streptococcus pneumoniae in search for potential common pneumococcal proteins as vaccine antigens. mAb 230,B-9 (IgG1) reacted by immunoblotting with a 70-kDa protein which was isolated by immunoaffinity chromatography and subsequent preparative electrophoresis. N-terminal amino acid sequencing showed homology to that of heat shock protein 70 (hsp70). The hsp70 epitope reactive with mAb 230,B-9 was found in all the pneumococci examined as well as in other streptococci and enterococci. The epitope was not expressed in several other examined Gram-positive or -negative bacteria. Pneumococcal hsp70 has by other investigators been proposed to be a vaccine candidate. Binding experiments using flow cytometry showed that the epitope was not surface-exposed on live exponential phase grown S. pneumoniae. Human patient sera did not react with affinity-purified pneumococcal hsp70. Therefore the pneumococcal hsp70 does not seem to be of special interest in a vaccine formulation. The human sera contained antibodies to high molecular proteins co-purified with hsp70. Some of these proteins could be the pneumococcal surface protein A.

show abstract

Monoclonal antibodies with specificities for Streptococcus pneumoniae group 9 capsular polysaccharides

Cited by 5 publications

References 13 publications

Streptococcus pneumoniae enolase is important for plasminogen binding despite low abundance of enolase protein on the bacterial cell surface

Streptococcus pneumoniae enolase is important for plasminogen binding despite low abundance of enolase protein on the bacterial cell surface

Epitope mapping of pneumococcal surface protein A of strain Rx1 using monoclonal antibodies and molecular structure modelling

Streptococcus pneumoniaeheat shock protein 70 does not induce human antibody responses during infection

Contact Info

Product

Resources

About