Proteins released during high toxin production in Clostridium difficile

The mechanism by which toxins A and B are released by Clostridium difficile is unknown and information about the other extracellular proteins of this bacterium is limited. The authors identified exported proteins from C difficile strain VPI 10463 during conditions promoting high toxin production. Toxins A and 8 were released in a 1: 1 ratio and the proportion of toxin in the extracellular fraction reached 50% during the stationary phase as compared to a proportion of <1% for typical cytoplasmic proteins, showing that toxin export was not due to bacterial lysis. A 47 kDa protein, released with similar kinetics to the toxins, was processed and showed weak similarity to the channel-forming protein TolC. Another protein released during high toxin production was unprocessed and showed similarity to XkdK encoded by the prophage PBSX in Bacillus subtilis, a protein supposedly exported via phage-specific holins. The two most abundant extracellular C difficile proteins, found during both high and low toxin production, were processed and identified as shed 5-layer proteins. As shown by N-terminal sequencing and PCR-based methods, there was a considerable sequence variation of the S-layer gene slpA in different serogroup reference strains. To conclude, C. difficile uses the classical Sec-dependent and probably also holin-like pathways to secrete a comparatively small repertoire of proteins.