Previous studies have shown that the detection of
antibodies to an 18-kDa cytoplasmic
protein of Brucella spp. is useful for the diagnosis of human and animal
brucellosis. This
protein has now been expressed in recombinant form in Escherichia coli. The
recombinant protein is soluble only under reducing conditions, but alkylation with
iodoacetamide renders it soluble in non-reducing media. As shown by gel exclusion chromatography, this soluble form arranges in pentamers of 90 kDa. The reactivity of human and animal sera against the
recombinant protein was similar to that found with the native
protein present in brucella cytoplasmic fraction, suggesting that the
recombinant protein is correctly folded. The
protein has low but significant homology (30%) with
lumazine synthases involved in bacterial
riboflavin biosynthesis, which also arrange as pentamers.
Biological tests on the
crude extract of the recombinant bacteria and on the purified
recombinant protein showed that the
biological activity of the Brucella spp. 18-kDa
protein is that of
lumazine synthase. Preliminary crystallographic analysis showed that the Brucella spp.
lumazine synthase arranges in icosahedric capsids similar to those formed by the
lumazine synthases of other bacteria. The high immunogenicity of this
protein, potentially useful for the design of
acellular vaccines, could be explained by this polymeric arrangement.