title: A novel 3-deoxy-D-manno-octulosonic acid (Kdo) hydrolase that removes the outer Kdo sugar of Helicobacter pylori lipopolysaccharide.
abstract: The lipid A domain anchors lipopolysaccharide (LPS) to the outer membrane and is typically a disaccharide of glucosamine that is both acylated and phosphorylated. The core and O-antigen carbohydrate domains are linked to the lipid A moiety through the eight-carbon sugar 3-deoxy-D-manno-octulosonic acid known as Kdo. Helicobacter pylori LPS has been characterized as having a single Kdo residue attached to lipid A, predicting in vivo a monofunctional Kdo transferase (WaaA). However, using an in vitro assay system we demonstrate that H. pylori WaaA is a bifunctional enzyme transferring two Kdo sugars to the tetra-acylated lipid A precursor lipid IV(A). In the present work we report the discovery of a Kdo hydrolase in membranes of H. pylori capable of removing the outer Kdo sugar from Kdo2-lipid A. Enzymatic removal of the Kdo group was dependent upon prior removal of the 1-phosphate group from the lipid A domain, and mass spectrometric analysis of the reaction product confirmed the enzymatic removal of a single Kdo residue by the Kdo-trimming enzyme. This is the first characterization of a Kdo hydrolase involved in the modification of gram-negative bacterial LPS.
authors.affiliation: Department of Microbiology, J. H. Quillen College of Medicine, Johnson City, Tennessee 37614, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Stead
authors.forename: An
authors.initials: A
authors.lastname: Tran
authors.forename: Donald
authors.initials: D
authors.lastname: Ferguson
authors.forename: Sara
authors.initials: S
authors.lastname: McGrath
authors.forename: Robert
authors.initials: R
authors.lastname: Cotter
authors.forename: Stephen
authors.initials: S
authors.lastname: Trent
mesh_terms: D016480:Helicobacter pylori / Q000201:enzymology* / Q000235:genetics; D006867:Hydrolases / Q000235:genetics / Q000378:metabolism*; D008050:Lipid A / Q000378:metabolism; D008070:Lipopolysaccharides / Q000737:chemistry / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013400:Sugar Acids / Q000737:chemistry / Q000378:metabolism*; D013696:Temperature; D014166:Transferases / Q000235:genetics / Q000378:metabolism
chemical_list: D008050:Lipid A; D008070:Lipopolysaccharides; D010710:Phosphates; D013400:Sugar Acids; C002532:2-keto-3-deoxyoctonate; D014166:Transferases; C019917:3-deoxy-D-manno-octulosonate transferase; D006867:Hydrolases