PhD ceremony Ms. L. Krishnappa: Proteolysis of extracytoplasmic proteins in Bacillus subtilis

PhD ceremony: Ms. L. Krishnappa, 14.30 uur, Academiegebouw, Broerstraat 5, Groningen

Dissertation: Proteolysis of extracytoplasmic proteins in Bacillus subtilis

Promotor(s): prof. J.M. van Dijl

Faculty: Medical Sciences

All living organisms have a limited lifespan and this is also true for their individual components. This thesis describes how proteolytic enzymes determine the 'life and death' of proteins that bacteria export from their site of synthesis - the cytoplasm - to the cytoplasmic membrane, the cell wall and the extracellular environment. For this purpose, the Gram-positive bacterium B. subtilis, a well-known 'cell factory' for secreted enzymes of high commercial value, was used as a model organism. Importantly, B. subtilis produces at least ten major extracytoplasmic proteases. Two of these are located in the membrane, one is located in the cell wall, and seven others are secreted into the extracellular milieu. Previous studies showed that several extracytoplasmic proteases set limits to the biotechnological use of B. subtilis, but it was not known to what extent these proteases degrade the bacterium's own proteins. Remarkably, the present investigations show that proteins in the membrane, cell wall and extracellular milieu are subject to extensive proteolysis. Unexpectedly, several proteases also attack components of the protein secretion machinery of B. subtilis, and this explains at least in part why multiple protease mutant cells have a generally improved capacity for protein secretion. On the other hand, several extracytoplasmic proteins seem to require certain proteases for their stable production. This is consistent with the view that some of the investigated proteases have important quality control functions. Altogether, these novel insights lay the foundation for the construction of next-generation super-secreting bacterial cell factories.