BIOCHEMISTRY, MOLECULAR & CELL BIOLOGY FIELD SEMINAR –
Kelly Rosch –
Doerr Lab –
Wednesday, October 11, 2023 –
2:55 PM | 105 Riley-Robb Hall –
Committee: Linda Nicholson, Joe Peters –
Vibrio cholerae endopeptidase ShyA mediates adaptation to osmotic stress via its LysM domain – 
The bacterial cell wall is a covalently closed meshwork that protects cells from osmotic lysis and maintains cell shape. The cell wall is made of peptidoglycan (PG), which consists of polysaccharide strands that are connected via short peptide crosslinks. As bacteria elongate and divide, PG is constantly remodeled through the activity of both cell wall synthases and hydrolases. PG hydrolysis is inherently dangerous to the cell, as unregulated PG cleavage activity results in cell lysis. Therefore, synthesis and hydrolysis are presumably tightly coupled to prevent structural integrity failure; but how this balance is maintained remains poorly understood. One such PG hydrolase, the endopeptidase ShyA, is regulated through conformational switching between an open (active) conformation and a closed (inactive) conformation. Despite exhibiting potent endopeptidase activity, ShyA does not cause cell lysis when overexpressed. These observations led us to hypothesize that ShyA is produced in the closed (inactive) conformation, and is activated by local PG stretching. To investigate this, I targeted potential PG-binding residues in the LysM carbohydrate-binding domain using site-directed mutagenesis and assessed cell viability and morphology on salt-free medium, where we expect enhanced PG stretching due to increased turgor pressure. Indeed, the LysM domain of ShyA is required for adaptation to osmotic stress, consistent with a model where PG binding is required for optimal ShyA function in fluctuating environments.