Kenneth Hatter

STRUCTURAL ANALYSIS AND PROTEOLYTIC ACTIVATION OF ENTEROCOCCUS FAECALIS CYTOLYSIN, A NOVEL LANTIBIOTIC

Clinical isolates of Enterococcus faecalis more commonly produce a cytolysin than do commensal isolates. Epidemiologic evidence and animal‐model studies have established a role for the cytolysin in the pathogenesis of enterococcal disease. The cytolysin consists of two structural subunits, CylLL and CylLs, that are activated by a third component, CylA. Genetic and biochemical characterization of CylA indicate that it is a serine protease, and that activation putatively results from cleavage of one or both cytolysin subunits. Genetic evidence also suggests that the cytolysin subunits are related to the rapidly growing class of bacteriocins termed lantibiotics. However, unlike lantibiotics, the cytolysin is lytic for eukaryotic as well as prokaryotic cells, and it consists of two structural subunits. This report describes the purification and characterization of the cytolysin subunits and detection of lanthionine‐type post‐translational modifications within their structures. Furthermore, the cleavage specificity of the CylA activator is reported and it is shown that proteolytic activation of both subunits is essential for activity.

Growth, development, and gene expression in a persistent Streptococcus gordonii biofilm.

ABSTRACT A model for the protracted (30-day) colonization of smooth surfaces by Streptococcus gordonii that incorporates the nutrient flux that occurs in the oral cavity was developed. This model was used to characterize the biphasic expansion of the adherent bacterial population, which corresponded with the emergence of higher-order architectures characteristic of biofilms. Biofilm formation by S. gordonii was observed to be influenced by the presence of simple sugars including sucrose, glucose, and fructose. Real-time PCR was used to quantify changes in expression of S. gordonii genes known or thought to be involved in biofilm formation. Morphological changes were accompanied by a significant shift in gene expression patterns. The majority of S. gordonii genes examined were observed to be downregulated in the biofilm phase. Genes found to be upregulated in the biofilm state were observed to encode products related to environmental sensing and signaling.