Cloning and expression of the porA gene of the Neisseria meningitidis strain B : 4 : P1.15 in Escherichia coli. Preliminary characterization of the recombinant polypeptide

Abstract

AbstractThe gene coding for the class 1 outer membrane protein from the Neisseria meningitidis strain B385 (B : 4 : P1.15) was isolated by the Polymerase Chain Reaction (PCR) and cloned into the Sma I cut M13mp 18 vector. Then, a Xba I restriction site was created by using an oligonucleotide‐directed in vitro mutagenesis system at the start of the coding fragment. In order to express the protein, this fragment was fused to 180 base pairs corresponding to 60 amino acids from the N terminus of interleukin‐2 under the control of the tryptophan promoter (Ptrp). The expression was confirmed by Western‐blotting where the recombinant protein (PILM28) was detected by bactericidal monoclonal antibodies (MABs). The recombinant polypeptide was partially purified and used to elicit murine antibodies, able to recognize the meningococcal class 1 subtype 15.