Molecular and Immunological Characterization of Glyceraldehyde-3-phosphate Dehydrogenase (GAPDH-1) of Neisseria meningitidis serogroup B strain MC58

Abstract

The present study focuses on characterizing the putative roles of GapA-1with aim to determine the additional non glycolytic roles of the enzyme in vaccine development and pathogenesis of N. meningitidis. To achieve the goals, a N. meningitidis GapA-1 isogenic knock-out mutant constructed previously was used to serve as a negative control in characterisation experiments. Sub-cellular localisation of GapA-1 was investigated by using cell fractionation method. The DNA fragment coding for GapA1 gene was amplified from a panel of 18 meningococcal clinical isolates by PCR. Strains yielding positive results for GapA-1 gene by PCR were further investigated for the expression of GapA-1 gene by western blot analysis using rabbit polyclonal antiserum (R?GAPDH-1). Furthermore, to assess the vaccine potential of GapA-1, in vitro serum bactericidal assay (BSA) was conducted using polyclonal R?GAPDH-1 antiserum. The gene encoding GapA1 was present in all strains and also expressed under in vitro conditions tested. However, GAPDH-1 antiserum raised against denatured purified protein failed to kill meningococci in vitro BSA experiments. In summary, GapA-1, is well conserved, constitutively expressed protein and localized to both cytoplasm and to the cell surface of Neisseria meningitidis serogroup B strain MC-58.