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Cloning, nucleotide sequence, and expression of the gene coding for a ribosome releasing factor-homologous protein of
N Vizcaíno1, A Cloeckaert, G Dubray
1Laboratoire de Pathologie Infectieuse et Immunologie, Centre de Recherches de Tours, Institut National de la Recherche Agronomique, Nouzilly, France. Nieves.Vizcaino@tours.inra.fr.
Abstract:
The gene coding for a Brucella melitensis cytosoluble protein (CP24) that is immunogenic in infected sheep and a major component of brucellin INRA was cloned and sequenced. As in Brucella cells, CP24 was located in the cytoplasm of recombinant Escherichia coli. The amino acid sequence predicted from the cloned gene revealed 48 and 46% identity with the ribosome releasing factor, a protein factor required for release of the 70S ribosome from the mRNA, of E. coli and Haemophilus influenzae Rd, respectively. Sera from naturally infected sheep and sheep experimentally infected with B. melitensis H38 showed antibody reactivity against recombinant CP24.
Insights
Brucella melitensis cytosoluble protein (CP24) was cloned and sequenced. This protein is immunogenic in sheep and shares similarities with bacterial ribosome releasing factors.
Area of Science:
- Microbiology
- Immunology
- Molecular Biology
Background:
- Brucella melitensis is a pathogen causing brucellosis in livestock and humans.
- Brucellin INRA is a diagnostic antigen derived from B. melitensis.
- A specific cytosoluble protein (CP24) is a major component of brucellin and is immunogenic.
Purpose of the Study:
- To clone and sequence the gene encoding the Brucella melitensis cytosoluble protein (CP24).
- To investigate the localization and potential function of CP24.
- To assess the immunogenicity of CP24 in infected sheep.
Main Methods:
- Gene cloning and sequencing of the CP24 gene from Brucella melitensis.
- Expression of recombinant CP24 in Escherichia coli to determine its subcellular localization.
- Bioinformatic analysis of the predicted amino acid sequence of CP24.
- Serological testing of sheep sera against recombinant CP24.
Main Results:
- The gene for CP24 was successfully cloned and sequenced.
- Recombinant CP24 was found to be localized in the cytoplasm of Escherichia coli, similar to its location in Brucella.
- The amino acid sequence of CP24 showed significant homology (48% and 46%) to ribosome releasing factors from E. coli and Haemophilus influenzae Rd.
- Antibodies against recombinant CP24 were detected in sera from both naturally and experimentally infected sheep.
Conclusions:
- CP24 is an immunogenic protein of Brucella melitensis.
- CP24's sequence homology suggests a potential role in protein synthesis or ribosome regulation.
- Recombinant CP24 is a valuable candidate antigen for diagnosing Brucella infections in sheep.