title: Revised mechanism of D-alanine incorporation into cell wall polymers in Gram-positive bacteria.
abstract: Teichoic acids (TAs) are important for growth, biofilm formation, adhesion and virulence of Gram-positive bacterial pathogens. The chemical structures of the TAs vary between bacteria, though they typically consist of zwitterionic polymers that are anchored to either the peptidoglycan layer as in the case of wall teichoic acid (WTA) or the cell membrane and named lipoteichoic acid (LTA). The polymers are modified with D-alanines and a lack of this decoration leads to increased susceptibility to cationic antimicrobial peptides. Four proteins, DltA-D, are essential for the incorporation of d-alanines into cell wall polymers and it has been established that DltA transfers D-alanines in the cytoplasm of the cell onto the carrier protein DltC. However, two conflicting models have been proposed for the remainder of the mechanism. Using a cellular protein localization and membrane topology analysis, we show here that DltC does not traverse the membrane and that DltD is anchored to the outside of the cell. These data are in agreement with the originally proposed model for D-alanine incorporation through a process that has been proposed to proceed via a D-alanine undecaprenyl phosphate membrane intermediate. Furthermore, we found that WTA isolated from a Staphylococcus aureus strain lacking LTA contains only a small amount of D-alanine, indicating that LTA has a role, either direct or indirect, in the efficient D-alanine incorporation into WTA in living cells.
authors.affiliation: Section of Microbiology and MRC Centre for Molecular Bacteriology and Infection, Imperial College London, London SW7 2AZ, UK.
authors.forename: Nathalie T
authors.initials: NT
authors.lastname: Reichmann
authors.affiliation: Section of Microbiology and MRC Centre for Molecular Bacteriology and Infection, Imperial College London, London SW7 2AZ, UK.
authors.forename: Carolina Picarra
authors.initials: CP
authors.lastname: Cassona
authors.affiliation: Section of Microbiology and MRC Centre for Molecular Bacteriology and Infection, Imperial College London, London SW7 2AZ, UK.
authors.forename: Angelika
authors.initials: A
authors.lastname: Gründling
mesh_terms: D000409:Alanine / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001704:Biopolymers / Q000737:chemistry / Q000378:metabolism*; D002473:Cell Wall / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000737:chemistry / Q000378:metabolism; D013211:Staphylococcus aureus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013682:Teichoic Acids / Q000737:chemistry / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D001704:Biopolymers; D008070:Lipopolysaccharides; D013682:Teichoic Acids; C009900:lipoteichoic acid; D000409:Alanine