Asunto(s)
Proteínas de la Membrana Bacteriana Externa/química , Proteínas de la Membrana Bacteriana Externa/inmunología , Klebsiella pneumoniae/química , Klebsiella pneumoniae/inmunología , Péptidos/química , Péptidos/inmunología , Polisacáridos Bacterianos/química , Polisacáridos Bacterianos/inmunología , Vacunas Conjugadas/química , Vacunas Conjugadas/inmunología , Animales , Proteínas Portadoras/química , Fenómenos Químicos , Química Física , Ensayo de Inmunoadsorción Enzimática , Escherichia coli/metabolismo , Inmunoglobulina G/biosíntesis , Inmunoglobulina G/química , Ratones , Ratones Endogámicos BALB CRESUMEN
Klebsiella pneumoniae OmpA, the 40-kDa major protein of the outer membrane, was cloned and expressed in Escherichia coli. The recombinant protein was produced intracellularly in E. coli as inclusion bodies. Fusion of a short peptide to the N-terminus of native P40 facilitated high-level expression of the recombinant protein. Purified recombinant P40 was analyzed to verify purity and structural integrity. The molecular mass of purified recombinant P40 determined by electrospray mass spectrometry was 37,061 Da, in agreement with the theoretical mass deduced from the DNA sequence. Specific proliferation of recombinant-P40-primed murine lymph node cells in response to recombinant P40 stimulation in vitro indicated the presence of a T-cell epitope on recombinant P40. The induction of high serum antibody titers to a synthetic peptide derived from the attachment protein G of the respiratory syncytial virus when chemically coupled to recombinant P40 indicated that the protein had potent carrier properties.