Self-assembled particulate PsaA as vaccine against Streptococcus pneumoniae infection

Abstract

Streptococcus pneumoniae is a human pathogen responsible for the majority of childhood pneumonia and media otitis cases worldwide. The diversity of its capsular polysaccharides (CPS) results in more than 91 serotypes of which at least 23 are virulent. Various CPS conjugated to immunogenic carrier proteins are currently licensed and provide protection against the infection caused by the respective serotypes but not against new and emerging virulent serotypes. In this study, we considered the conserved protein antigen PsaA, the pneumococcal surface adhesin A, in order to overcome the limitations of CPS antigens. The PsaA was translationally fused to a polyhydroxybutyrate (PHB) synthase which mediated production of PsaA displayed on PHB inclusions in recombinant Escherichia coli. This suggested that the PsaA fusion to the PHB synthase did not interfere with PHB synthase activity and its ability to mediate formation of nano-sized inclusions composed of a PHB core surrounded by the PHB synthase fused to PsaA. Isolated PHB beads showed a negative surface charge. Transmission electron microscopy analysis suggested that the PsaA fusion to the PHB synthase reduced the size of PHB beads from about 500 nm to 100 nm. The integrity and antigenicity of the fusion protein attached to isolated PHB beads was confirmed by SDS-PAGE, tryptic peptide fingerprinting analysis using MALDI-TOF-MS/MS and immunoblotting using a monoclonal anti-PsaA antibody. Mice immunized with PsaA displaying PHB beads produced high and specific IgG levels dominated by IgG1 isotype. While IgG1 titer were similar between soluble and insoluble PsaA, the IgG2 titers were strongly increased upon vaccination with insoluble PsaA i.e. PsaA displayed on PHB beads. Particulate PsaA-PHB beads elicited IgG antibodies recognizing PsaA in whole cell lysates of seven different serotypes of S. pneumoniae. This study suggested that PHB beads are suitable carriers for PsaA in order to induce a significant and specific Th-2-type immune response.

Bioengineered polyester beads co-displaying protein and carbohydrate-based antigens induce protective immunity against bacterial infection

“…Ratio ng of the fusion protein/ mg of the wet bead was determined as mentioned above 80 . The protein concentrations of NadA-His6 and GNA2091-fHbp-G1-His6 were determined by BCA colorimetric assay (Pierce™ BCA Protein Assay Kit) 81 .…”

Covalent Functionalization of Bioengineered Polyhydroxyalkanoate Spheres Directed by Specific Protein-Protein Interactions

“…In addition, WT -displaying PHA spheres and their recombinant variants often exhibited a negative surface charge at pH 7.5. The isoelectric point (pI) of PHA spheres in suspension typically varies between approximately 5-6 (González-Miro et al, 2017;Gonzalez-Miro et al, 2018a,b). However, the predicted pI of the SdyTag peptide itself is pH 3.9, much lower than those compared to both SpyTag and SnoopTag at pH 8.6 and pH 8.5, respectively (Gasteiger et al, 2005).…”

Next-Generation Whole-Cell Pneumococcal Vaccine

“…The first protein antigens were identified through antibody screening, and specific antibodies were found in children exposed to pneumococcus, namely pneumolysin (Ply), pneumococcal surface protein A (PspA), pneumococcal surface protein C (PspC), and pneumococcal surface antigen A (PsaA) [25]. These proteins were proposed for use in a vaccine with the capacity to induce serotype-independent antibodies and broader protective immunity against pneumococcal infections [31,32].…”

“…PsaA is a conserved lipoprotein that transports Mn 2+ and also functions as adhesin, which plays a major role in pneumococcal attachment to the host cell and virulence [30,32].…”