The bacterial cell wall is a complex structure that exerts essential protective functions against environmental insults, allowing bacterial adaptation under adverse conditions. Biopolymers, such as teichoic acids and capsular polysaccharides, play fundamental roles in immune evasion, adhesion, biofilm formation, and protection against antimicrobials. Our goal is to reveal the molecular and mechanistic basis of cell wall assembly. How does the cell acquire, synthesize, and move essential precursor molecules? How does the polymerization of cell wall components occur?  Can we engineer and use to our advantage the proteins and pathways involved in cell wall synthesis? To address these questions, we use single-particle cryo-EM, X-ray crystallography, Solid-Supported-Membranes (SSM) electrophysiology, multiple biochemical and biophysical technique for the study of membrane protein mechanisms, in vitro bacterial assays, and nanobodies.

The biosynthesis of cell wall biopolymers generally requires the transport of lipids and soluble precursors across the plasma membrane of bacteria, involving the participation of active and passive transporters and flippases. The mechanisms of these proteins are not well understood. In the last years, our Lab has investigated the mechanism, activity, specificity, and inhibition of transporters and flippases participating in teichoic acid synthesis in two prominent Gram-positive bacteria pathogens, Staphylococcus aureus and Streptococcus pneumoniae. We elucidated structures of fundamental conformational states of these proteins and described previously unknown architectural features essential to understanding their mechanisms. The essential role of transporters in cell wall biosynthesis makes them valuable drug targets. Our Lab is working on establishing a framework for generating transporters inhibitors based on nanobodies and small molecules.

Bacterial cells can be subjected to environmental changes. How do they adapt to challenging conditions? How does the cell wall contribute to adaptation? What are the molecular and mechanistic basis of these adaptation processes? Recently we revealed that LtaA is a proton-coupled lipid flippase essential to combat acidic stress in the pathogen Staphylococcus aureus. In S. aureus, LTA synthesis initiates at the cytoplasmic leaflet of the plasma membrane by the assembly of an anchor glycolipid, a step catalyzed by the membrane-associated glycosyltransferase YpfP. The anchor glycolipid is later translocated across the membrane by the flippase LtaA. This translocation process is essential for the LtaS-mediated lipoteichoic acid synthesis and constitutes a rate-limiting step in the biosynthesis pathway. We have revealed that LtaA is a “pH sensing” flippase that contributes to adjusting the pool of lipoteichoic acids in the cell wall helping S. aureus to withstand host niche pH conditions i.e. the human nasopharynx, mucous membranes, and skin, which present mild acidic environments (5.0<pH<6.5). Our lab is interested in revealing the role of this and other transporters in bacterial pathogens adaptation.