Mycoplasma hyopneumoniae is an economically significant pathogenic bacterium that chronically infects the respiratory tract of swine. This infection leads to pneumonia and secondary infections, costing agricultural industries significantly in the use of antibiotics and vaccines, which are currently largely ineffective. An improved understanding of the molecular mechanisms behind the infection process is essential to our ability to rationally design better vaccine and therapeutic interventions. With fewer than 700 predicted protein coding sequences, M. hyopneumoniae possesses one of the smallest genomes of any free-living organism. As such, it lends itself well to thorough proteomic interrogation.

A slew of proteomic techniques have been used to investigate the global proteome and surface-proteome at the protein and peptide level, including surface shaving and labelling techniques, ligand and immuno-blotting and affinity chromatography, as well as N-terminal dimethyl labelling to determine true N-termini of mature proteins.

This conceptually unbiased, function-oriented approach has revealed an unexpected level of complexity in the use of proteolytic processing, multifunctional proteins and moonlighting to compensate for reduced coding capacity at the genome level. While transcriptome studies suggest that the majority of genes are transcribed, our analyses identified less than 400 detectable expressed protein products. However, a significant number of these were discovered to be post-translationally modified, multifunctional or present at the cell surface, despite not possessing traditional signal peptides. Many of these proteins, predominantly cell surface adhesins, we have already described in the literature, however a surprising number of cytoplasmic “housekeeping” proteins are also found to be post-translationally cleaved, multifunctional or moonlighters. These findings can be applied to the development of vaccines and therapeutics in Mycoplasma, as well as having wider implications for the field of biology, if this level of post-translational regulation can be found in other bacterial pathogens.