Sequence, cloning, expression and characterisation of the 81-kDa surface membrane protein (P80) of Mycoplasma
1Istituto Zooprofilattico Sperimentale della Sardegna "G. Pegreffi", Sassari, Italy. sebastiana.tola@tin.it
Abstract:
Mycoplasma agalactiae, the causative agent of contagious agalactia in small ruminants, produces a protein, named P80, that is detectable in all wild-type isolates examined to date and that appears expressed during the early phase of infection. We describe here the identification, cloning and expression of the gene encoding P80 (ma-mp81). The deduced amino acid sequence is consistent with a hydrophobic and basic protein that possesses a lipoprotein signal peptide. Sequence analysis of gene ma-mp81 suggests that P80 is a membrane lipoprotein that shows significant homology with other putative lipoproteins of M. pneumoniae. An internal 1-kb fragment of ma-mp81 was expressed in Escherichia coli as a 6xHis-tagged protein. The purified recombinant protein greatly reacted with polyclonal anti-P80 sera raised in lamb.
Insights
Mycoplasma agalactiae produces a P80 protein during early infection in small ruminants. Researchers identified, cloned, and expressed the gene encoding this membrane lipoprotein, ma-mp81.
Area of Science:
- Veterinary Microbiology
- Molecular Biology
- Protein Biochemistry
Background:
- Mycoplasma agalactiae causes contagious agalactia in small ruminants.
- A protein, P80, is detected in all wild-type isolates and expressed early in infection.
Purpose of the Study:
- To identify, clone, and express the gene encoding the P80 protein (ma-mp81).
- To characterize the P80 protein's sequence and potential function.
Main Methods:
- Gene identification and cloning of ma-mp81.
- Expression of an internal ma-mp81 fragment in Escherichia coli as a 6xHis-tagged protein.
- Purification of the recombinant protein.
Main Results:
- The deduced amino acid sequence indicates P80 is a hydrophobic, basic lipoprotein with a signal peptide.
- Sequence analysis reveals homology with Mycoplasma pneumoniae lipoproteins.
- The purified recombinant P80 protein reacted strongly with anti-P80 sera.
Conclusions:
- The P80 protein is a membrane lipoprotein encoded by the ma-mp81 gene.
- The recombinant P80 protein is immunogenic and can be used for diagnostic purposes.
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