FUNCTIONAL ANALYSIS OF THE LACTOCOCCAL Bce-LIKE COMPLEX INVOLVED IN CELL WALL DAMAGE IN Lactococcus lactis

Lactococcus lactis, a model bacterium within lactic acid bacteria (LAB), is widely used as a starter in the manufacture of fermented dairy products. It’s an industrially relevant bacteria that has been thoroughly studied and is becoming the host of choice for multiple biotechnological applications as a cell factory. L. lactis is exposed to stressful conditions that compromise its viability, whether used as a dairy starter or cell factory. The majority of stressful situations that L. lactis encounters during industrial operations have an effect on the cell wall. To address the demand for robust strains, the DairySafe research team has concentrated on understanding how this bacteria detects and reacts to damage to the cell envelope. L. lactis deploys different pathways to detect cell wall damage and produce a response that triggers defence mechanisms. One of these sensing devices is the Bce-like sensory complex involving a two-component system (TCS-G) and an ABC transporter that detects specifically cell wall-acting antimicrobial peptides and provides resistance to them. Moreover, the presence of accessory proteins, a transmembrane protein YsaA of unknown function and a putative secreted peptide YsaD, makes the lactococcal Bce-like sensory complex distinctive. The goal of this thesis work is to carry out a functional analysis of the lactococcal Bce-like complex and the genes that are under its control. In-frame deletions have been introduced by homologous recombination for the inactivation of the genes of the additional proteins ysaD and ysaA that characterize the L. lactis Bce-like module to evaluate their impact on signal transduction and, thereby, on the cell response. The findings show whether signal transduction is lost or changed when any of these proteins are absent.