Neisseria meningitidis strains grown under
iron starvation conditions produce
transferrin binding proteins (Tbp1 and Tbp2) which have been shown to play a major role in
iron acquisition. Recent studies performed with Tbp2 purified from N. meningitidis suggest that this
surface protein is a potential
vaccine component. In order to further evaluate the immunogenicity of Tbp2, it was essential to develop a heterologous expression system to generate high amounts of purified
protein. Tbp2 is produced in Neisseria as a precursor with a
signal peptide whose cleavage follows a lipidation step on a
cysteine residue which is the first
amino acid in the mature
protein. When produced in Escherichia coli with its natural
signal peptide, a high amount of Tbp2 (about 10% of total cell
proteins) was detected. However, most of the
protein was nonlipidated precursor and only a small fraction was mature Tbp2. In order to optimize the maturation of the precursor, the natural
signal sequence was replaced by several E. coli
lipoprotein signal peptides. Expression levels and maturation of the precursor were highly variable depending on the
signal peptide used. With one of these, an efficient maturation and a high amount of mature lipidated Tbp2 were obtained (about 3% of total cell
proteins). A large-scale production process was then established for this E. coli-produced Tbp2.