The 18-kDa cytoplasmic protein of Brucella species --an antigen useful for diagnosis--is a lumazine synthase.
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.
Goldbaum
Fernando A
FA
Instituto de Estudios de la Inmunidad Humoral (IDEHU), Facultad de Farmacia y Bioquímica UBA, Junin 956 4to. piso, 1113 Buenos Aires, Argentina.
Velikovsky
Carlos A
CA
Instituto de Estudios de la Inmunidad Humoral (IDEHU), Facultad de Farmacia y Bioquímica UBA, Junin 956 4to. piso, 1113 Buenos Aires, Argentina.
Baldi
Pablo C
PC
Instituto de Estudios de la Inmunidad Humoral (IDEHU), Facultad de Farmacia y Bioquímica UBA, Junin 956 4to. piso, 1113 Buenos Aires, Argentina.
Mörtl
Simone
S
Department of Chemistry, Technical University of Munich, Lichtenbergstrasse 4, D-85747, Garching, Germany.
Bacher
Adelbert
A
Department of Chemistry, Technical University of Munich, Lichtenbergstrasse 4, D-85747, Garching, Germany.
Fossati
Carlos A
CA
Instituto de Estudios de la Inmunidad Humoral (IDEHU), Facultad de Farmacia y Bioquímica UBA, Junin 956 4to. piso, 1113 Buenos Aires, Argentina.
D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D000942:Antigens, Bacterial / Q000276:immunology; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D002002:Brucella / Q000201:enzymology* / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D008074:Lipoproteins; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D008074:Lipoproteins; D009097:Multienzyme Complexes; C097319:OMP19 protein, Brucella abortus; D011994:Recombinant Proteins; C044822:6,7-dimethyl-8-ribityllumazine synthase