Regulation and function of the cell wall polymer lipoteichoic acid in staphylococcus aureus

A thesis submitted for the Degree of Master in Medical microbiology

Experimental work performed at the Centre for Molecular Bacteriology and Infection, Imperial College London

Lipoteichoic acid (LTA) is an important polymer present in the envelope of various Gram-positive bacteria. In Staphylococcus aureus it is composed of a polyglycerolphosphate chain synthesised by the enzyme LtaS, and is anchored to the membrane via a glycolipid anchor. Depletion of LTA is known to cause growth arrest, aberrant positioning of septa and enlargement of cells leading to eventual cell lysis. This highlights the importance of LTA for bacterial growth and also suggests a possible link between LTA synthesis and cell division. Although key enzymes required for LTA synthesis have been identified, how this process is regulated has not been elucidated. This study investigates the mechanisms of regulation of LTA synthesis, as well as provides further insights into the roles of LTA in S. aureus. In this study, nuclear magnetic resonance (NMR) was used to experimentally confirm a function for LTA in the incorporation of D-alanines into a second polymer, wall teichoic acid (WTA). LTA was also shown to be essential not only for bacterial growth, but also for bacterial survival, as cells become bactericidal in its absence. Furthermore, fluorescence microscopy using a GFP-tagged version of the synthase enzyme indicated that LtaS localises to the septum, suggesting a possible role for LTA in cell division. Regarding LTA regulation, this work has shown that the ltaS gene has two promoters and a large upstream UTR, but that only one promoter is essential and sufficient for expression. Studying the effect of salt stress on LTA production has also revealed that an increase the NaCl concentration in the media causes a reduction in the levels of LTA produced, which in turn has an effect on growth. However this is not due to a decrease in the levels of LtaS, reflecting a possible ionic or osmotic effect on LtaS enzymatic activity.