Extraction and Characterization of the Smooth‐Type Lipopolysaccharide from Fusobacterium nucleatum JCM 8532 and Its Biological Activities

Abstract

The lipopolysaccharide (LPS) of Fusobacterium nucleatum JCM 8532 was isolated by hot-phenol water extraction. Most of the LPS was extracted in the phenolic phase and shown to be the smooth-type, whereas the aqueous phase contained mainly rough-type LPS. The chemical composition of the LPS was similar to that reported in other studies, but D-quinovosamine, which may be a major component of O-antigenic polysaccharide, and 3-deoxy-D-manno-2-octulosonic acid (Kdo) were detected for the first time by gas chromatography-mass spectrometry. The biological activities of smooth-type LPS, including limulus activity, lethal toxicity, pyrogenicity, and B lymphocyte mitogenicity, were comparable to those of enterobacterial LPS. Smooth-type LPS inhibited the cell growth and DNA synthesis of adult and fetal human gingival fibroblasts in a dose-dependent manner, suggesting that LPS may play a role in the occurrence of human gingivitis.

Inducible Expression of Human β-Defensin 2 byFusobacterium nucleatumin Oral Epithelial Cells: Multiple Signaling Pathways and Role of Commensal Bacteria in Innate Immunity and the Epithelial Barrier

“…Purified LPS fraction of F. nucleatum ATCC 25586, grown in mycoplasma broth supplemented with hemin and menadione, was prepared by the cold magnesium-ethanol precipitation technique (11) followed by lipid extraction (16) and conversion to sodium salts (34). The LPS preparations were subjected to gas chromatography for analysis of carbohydrates (8) and fatty acids (43) and were found to have compositions consistent with a previous report (33) and to be devoid of phospholipids and procedure-related detergent. The A 280 and A 260 indicated the LPS preparation was free of detectable levels of protein and nucleic acid contamination.…”

Serum Antibodies of Periodontitis Patients Compared to the Lipopolysaccharides of Porphyromonas gingivalis and Fusobacterium nucleatum

“…The P-LPS contained glucose instead of galactose and a larger amount of D-quinovosamine compared with the A-LPS. Other components were similar in both LPS preparations, as described previously (15). Antibody titers against the LPSs of P. gingivalis and F. nucleatum were measured by enzyme-linked immunosorbent assay (ELISA) as described by Amano et al (1).…”

“…nucleatum JCM 8532 T was grown as described previously (15), and LPS was extracted by hot-phenol water extraction (25). The aqueous and phenolic phases were recovered and purified as described in a previous paper (15).…”

“…nucleatum JCM 8532 T was grown as described previously (15), and LPS was extracted by hot-phenol water extraction (25). The aqueous and phenolic phases were recovered and purified as described in a previous paper (15). The aqueous phase LPS (mainly roughtype LPS) was designated A-LPS and the phenolic phase LPS with long O-antigenic polysaccharide (smoothtype LPS) was designated P-LPS.…”

Soluble CD14 Discriminates Slight Structural Differences between Lipid As That Lead to Distinct Host Cell Activation

“…Bacterial cells were collected by centrifugation, then washed three times with saline, and lyophilized. The lyophilized cells were subjected to a hot-phenol water method, with the phenolic phase recovered to obtain a smooth-type LPS preparation, as previously described (22,24). Lipid A was then prepared from the LPS preparation according to a method described previously (25).…”