| Zusammenfassung |
Bacterial spores are among the most pernicious cells in nature. Formed as survival structures in response to nutrient starvation, their ubiquity and extreme resistance to heat and other stress factors that would ordinarily kill vegetative cells means they are difficult to eradicate. This presents problems in the food sector, for example, since spores can survive processing and then germinate and proliferate, resulting in spoilage or toxigenesis. Germination and ultimately improved control of spores is at the heart of the current proposal. In order to complete germination, spores must enzymatically degrade what is essentially their cell wall, a thick layer of peptidoglycan referred to as the cortex. This is mediated by cortex lytic enzymes, one of which – SleB – is somehow held in an inactive state during spore dormancy and which can withstand whatever insults are thrown at the spore during that period, before rapidly assuming activity during spore germination. The molecular mechanisms associated with regulation of SleB activity in dormant and germinating spores are largely unknown, but if understood should present new routes to improved control of spores. This project aims to solve the SleB puzzle while at the same time identifying and developing inhibitors of the enzyme. We will achieve this by: Combining X-ray crystallography and related techniques with Molecular Dynamics (MD) simulations to yield insight to the protein complex formed by SleB and its partner protein YpeB, revealing the molecular mechanisms that regulate the enzyme’s (in)activity during spore dormancy, exposure to heat, and germination. Solving the crystal structure of SleB with bound cortex peptidoglycan substrate, revealing the structural basis for substrate specificity by the enzyme. Conducting crystallographic fragment-based screens to identify small molecule ligands and potential inhibitors of SleB. Ultimately the project will fill crucial knowledge gaps in current models of spore germination while providing a platform for spore-associated interventions in global food production and nutrition, and in the public health, infectious disease, and animal welfare sectors. |
