• Biosynthesis of the Vibrio cholerae Kdo-lipid A Domain and its Role in Pathogenesis

      Hankins, Jessica V.; Department of Biochemistry and Molecular Biology (2011-05)
      Bacteria assemble remarkable surface structures that interface with their surrounding environment. One such structure is the glycolipid lipopolysaccharide (LPS) that covers the surface of Gram-negative bacteria. LPS is anchored to the bacterial cell by its lipid anchor known as lipid A. Since lipid A is the bioactive component of LPS, modulation of its structure can have a profound impact on disease by altering the host immune response. Additionally, LPS structure directly impacts the outer membrane permeability barrier and bacterial resistance to host antimicrobial peptides. Although the lipid A domain of Escherichia coli has been well characterized, the Vibrio cholerae lipid A biosynthetic pathway has received little attention. The late stages of lipid A biosynthesis include the transfer of the 3-deoxy-Dmanno- octulosonic acid (Kdo) sugars and the secondary acyl chains to the lipid A backbone. Here, the V. cholerae Kdo transferase (Vc0233) was shown to be monofunctional, transferring one Kdo residue to the lipid A precursor, lipid IVA. V. cholerae encode a Kdo kinase (Vc0227) responsible for the phosphorylation of the Kdo residue. The functionality of Vc0227 was shown to be required for the activity of the V. cholerae lipid A LpxL homologue, Vc0213. Interestingly, the addition of the phosphate group on the Kdo sugar was shown to be essential for lipid A secondary acylation in Haemophilus influenzae and Bordetella pertussis. Vc0213 was shown to catalyze the transfer of a myristate (C14:0) to the 2′-position of the V. cholerae phosphorylated Kdolipid A domain. A second protein, Vc0212, acts as an LpxM homologue and transfers 3- hydroxylaurate (3-OH C12:0) to the 3′-position creating hexa-acylated V. cholerae lipid A domain. Although lipid A secondary acyltransferases have been characterized among various Gram-negative bacteria, this is the first report of a lipid A secondary hydroxyacyltransferase. Further, the transfer of 3-hydroxylaurate (3-OH C12:0) was demonstrated to be essential for antimicrobial peptide resistance in V. cholerae and required for activation of the innate immune receptor TLR4.