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J. Bacteriol. doi:10.1128/JB.01868-07
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Identification of a Second Lipopolysaccharide in Porphyromonas gingivalis W50

Centre for Infectious Disease, Institute of Cell and Molecular Science, Centre for Clinical and Diagnostic Oral Sciences, Barts and The London Queen Mary's School of Medicine & Dentistry, 4 Newark Street, London E1 2AT, Medical Biomics Centre, St.George's University of London, Cranmer Terrace, London SW17 0RE

^* To whom correspondence should be addressed. Email: m.rangarajan{at}qmul.ac.uk.

	Abstract

We previously described a cell surface anionic polysaccharide (APS) in P.gingivalis which is required for cell integrity and serum resistance. APS is a phosphorylated-branched mannan which shares a common epitope with the post-translational additions to some of the Arg-gingipains. This study aimed to determine the mechanism of anchoring of APS to the surface of P.gingivalis. APS was purified on concanavalin A -affinity columns to minimise the loss of the anchoring system which occurred during chemical extraction. ¹H-NMR spectroscopy of the lectin-purified APS confirmed the previous structure but also revealed additional signals which suggested the presence of a lipid A. This was confirmed by fatty-acid analysis of the APS and MALDI-ToF MS of lipid A released by treatment with sodium acetate buffer pH4.5. Hence, P.gingivalis synthesises two distinct LPS macromolecules containing different glycan repeating units: O-LPS (with O-antigen tetrasaccharide repeating units) and A-LPS (with APS repeating units). Non-phosphorylated penta-acylated and non-phosphorylated tetra-acylated species were detected in the lipid A from P.gingivalis total LPS and in lipid A from A-LPS. These lipid A species were unique to lipid A derived from A-LPS. Biological assays demonstrated a reduced pro-inflammatory activity of A-LPS compared to total LPS. Inactivation of a putative O-antigen ligase (waaL) at PG1051, which is required for the final step of LPS biosynthesis abolished the linkage of both the O-antigen and APS to the lipid A-core of O-LPS and A-LPS respectively suggesting that WaaL in P.gingivalis has dual specificity for both O-antigen and APS repeating units.