Teichoic acids in the periplasm and cell envelope of Streptococcus pneumoniae

Abstract Teichoic acids (TA) are linear phospho-saccharidic polymers and important constituents of the cell envelope of Gram-positive bacteria, either bound to the peptidoglycan as wall teichoic acids (WTA) or to the membrane as lipoteichoic acids (LTA). The chemical composition of TA varies greatly but the presence of both WTA and LTA is highly conserved, hinting at an underlying fundamental function that is distinct from their numerous specific roles in diverse organisms. We report here the observation of a periplasmic space in the Gram-positive Streptococcus pneumoniae by cryo-electron microscopy of vitreous section. The thickness and appearance of this region change upon deletion of genes involved in the attachment of teichoic acids, supporting the role of TA in the maintenance of a periplasmic space in Gram-positive bacteria as their main universal function. Consequences of these mutations were further examined by super-resolved microscopy (dSTORM), following metabolic and fluorophore coupling by click-chemistry in pulse and pulse-chase experiments. This novel labeling method also enabled the titration the actual amount of TA per cell and to determine the ratio of WTA to LTA, to follow the change of TA length during growth phases, and to discover that a mutant devoid of LTA accumulates the membrane-bound TA precursor, that may compensate the absence of LTA. Most importantly, the possibility to follow TA during cell fractionation led to the discovery that LTA containing membranes sediment at low centrifugal forces, allowing easy separation.