Identification and biosynthesis of cyclic enterobacterial common antigen in Escherichia coli.
Phosphoglyceride-linked enterobacterial common antigen (ECA(PG)) is a cell surface glycolipid that is synthesized by all gram-negative enteric bacteria. The carbohydrate portion of ECA(PG) consists of linear heteropolysaccharide chains comprised of the trisaccharide repeat unit Fuc4NAc-ManNAcA-GlcNAc, where Fuc4NAc is 4-acetamido-4,6-dideoxy-D-galactose, ManNAcA is N-acetyl-D-mannosaminuronic acid, and GlcNAc is N-acetyl-D-glucosamine. The potential reducing terminal GlcNAc residue of each polysaccharide chain is linked via phosphodiester linkage to a phosphoglyceride aglycone. We demonstrate here the occurrence of a water-soluble cyclic form of enterobacterial common antigen, ECA(CYC), purified from Escherichia coli strains B and K-12 with solution nuclear magnetic resonance (NMR) spectroscopy, electrospray ionization mass spectrometry (ESI-MS), and additional biochemical methods. The ECA(CYC) molecules lacked an aglycone and contained four trisaccharide repeat units that were nonstoichiometrically substituted with up to four O-acetyl groups. ECA(CYC) was not detected in mutant strains that possessed null mutations in the wecA, wecF, and wecG genes of the wec gene cluster. These observations corroborate the structural data obtained by NMR and ESI-MS analyses and show for the first time that the trisaccharide repeat units of ECA(CYC) and ECA(PG) are assembled by a common biosynthetic pathway.
Erbel
Paul J A
PJ
Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9038, USA.
Barr
Kathleen
K
Gao
Ninguo
N
Gerwig
Gerrit J
GJ
Rick
Paul D
PD
Gardner
Kevin H
KH
D000942:Antigens, Bacterial / Q000096:biosynthesis* / Q000737:chemistry* / Q000302:isolation & purification; D051792:Basic Helix-Loop-Helix Proteins; D004586:Electrophoresis / Q000379:methods; D004755:Enterobacteriaceae / Q000276:immunology; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000254:growth & development / Q000276:immunology*; D009682:Magnetic Resonance Spectroscopy; D021241:Spectrometry, Mass, Electrospray Ionization; D015534:Trans-Activators / Q000302:isolation & purification / Q000378:metabolism; D000099162:Endothelial PAS Domain-Containing Protein 1
D000942:Antigens, Bacterial; D051792:Basic Helix-Loop-Helix Proteins; D015534:Trans-Activators; D000099162:Endothelial PAS Domain-Containing Protein 1; C017286:enterobacterial common antigen