Cloning and Expression of Pneumococcal Surface Protein A and Its Cross-immunoreactivity

Abstract:

In this study, we cloned genes coding the N-terminal region of PspA( pneumococcal surface protein A) from five of the most epidemic pneumoniae serotypes in China(FY01, FY05, FY6B, FY19F, FY23F). The obtained genes were respectively constructed into prokaryotic expression vector pET-27b(+), and then recombinant plasmids pET-pspA were respectively transformed into Escherichia coli BL21(DE3) for PspA expression. Induced with lactose, the five recombinant proteins were highly expressed and then purified by Ni-chelation chromatography. PCR with Family special primers was carried out to identify Family of PspA, and Clade of PspA was identified by means of bioinformatics according to the sequencing results. FY01rPspA, FY6B rPspA and FY19FrPspA belonged to Clade 1 of FamilyⅠ, FY05rPspA belonged to Clade 2 from FamilyⅠand FY23F rPspA belonged to Clade 3 from FamilyⅡ. Strong immunogenicity had been demonstrated in mice immuned with FY01rPspA at dose of 20 μg/mL each, of which antiserum antibody titer reached 1:210000. Result of western blotting showed that FY01rPspA antiserm reacted well with Y6B rPspAand FY19FrPspA but poorly with the other two, which suggested that cross-immunoreactivity was restricted in the same Clade. Mice immuned with FY01rPspA protected well against FY6B, while against FY05, FY23F was not effectively protected. The result in this study has much instructive significance for the development of a new efficient vaccine against pneumoniae.