Regulation of fructose metabolism and polymer synthesis by Fusobacterium nucleatum ATCC 10953

Abstract

Energy for the anaerobic growth of Fusobacterium nucleatum ATCC 10953 can be derived from the fermentation of sugar (fructose) or amino acid (glutamate). During growth on fructose, the cells formed large intracellular granules which after extraction yielded glucose by either acid or enzymatic hydrolysis. The endogenous polymer was subsequently metabolized, and after overnight incubation of the cells in buffer, the glucan granules were no longer detectable by electron microscopy. Anaerobically, washed cells grown previously on fructose fermented this sugar to a mixture of lactic, acetic, and butyric acids, and little intracellular glucan was formed. Aerobically, the cells slowly metabolized fructose to acetate. Fusobacterium nucleatum is a gram-negative, spindleshaped organism (5, 15, 17, 43) which by association (11,16,27,37) is believed to play a role in the etiology of periodontitis and gingivitis in humans (21,25,26,38). The bacterium relies primarily on the catabolism of amino acids (e.g., glutamate, histidine, lysine [2-4, 6, 10, 19, 23]) for energy production, and the end products of these fermentations are usually acetic, propionic, and butyric acids (4, 6, 17-19, 23, 44). Penetration of periodontal tissue by the cytotoxic organic acids (36), together with the amino acid-rich environment of the gingival crevice (24, 35), facilitate the invasion and provide an ecological niche conducive to establishment of the oral pathogen (11,16,27,38).Bergey's Manual of Systematic Bacteriology (18) and other sources (15,17,23) describe the fusobacteria as weakly saccharolytic. The early studies of Jackins and Barker (19), Loesche and Gibbons (23), and Coles (7), as well as more recent reports (9, 31), attest to the limited glycolytic activity of F. nucleatum. It is perhaps because of these observations that regulation of sugar transport and metabolism by F. nucleatum has received little attention. However, in the past 3 years (31-33), we have obtained partial explanations for the weakly saccharolytic capacity of this organism by the discoveries (i) that transport and phosphorylation of sugars (e.g., glucose and galactose) depend on ATP generated via amino acid metabolism, (ii) that intracellular sugar phosphates are rapidly converted to polymer, and (iii) that concomitant fermentation of amino acids can prevent the degradation of endogenous polymer. It seemed reasonable to assume that the transport and dissimilation of fructose would likewise depend on provision of a fermentable amino acid, but this proves not to be the case.This communication represents the first detailed descrip-* Corresponding author. tion of fructose transport and metabolism by F. nucleatum. In addition, our observations provide further insight into the unusual interrelationships which exist between amino acid and sugar fermentations in this organism (9,(31)(32)(33). Finally, we show how the cofermentation of an amino acid (glutamate) can regulate the entry of fructose into either the catabolic or polymer-synthesizing pathway of the cell. MATERIAL...

Sucrose fermentation by Fusobacterium mortiferum ATCC 25557: transport, catabolism, and products

“…At saturating concentrations (10 mM), fructose and mannose were phosphorylated at comparable rates, whereas 2-fluoro-mannose and mannosamine were phosphorylated at rates approximately 30 and 12% that of fructose, respectively. (45) revealed the formation of intracellular fructose 1-phosphate (the presumptive product of fructose-PEP:PTS activity [42]), attempts to demonstrate this group translocation system in either permeabilized cells or in reconstituted cell extracts were unsuccessful (45). To our knowledge, the present communication provides the first definitive evidence for PTS function in the Bacteroidaceae family in general and in the genus Fusobacterium specifically.…”

“…In contrast to the well-documented pathways for the metabolism of amino acids (e.g., glutamic acid [8] and lysine [3]), reports of sugar dissimilation by fusobacteria are few and controversial (10,22,32,43). Some of the controversial aspects may be explained by our recent discovery that, for most species, the transport, phosphorylation, and formation of an endogenous sugar polymer is dependent (or markedly enhanced) upon provision of a fermentable amino acid as the energy source (43)(44)(45) Harvesting and preparation of cells. Cells were harvested by centrifugation (13,000 x g for 10 min at 4°C), and the cell pellets were washed by resuspension and centrifugation from 50 mM potassium phosphate buffer (pH 7.0) containing 0.7 mM MgCl2.…”

Purification from Fusobacterium mortiferum ATCC 25557 of a 6-phosphoryl-O-alpha-D-glucopyranosyl:6-phosphoglucohydrolase that hydrolyzes maltose 6-phosphate and related phospho-alpha-D-glucosides

“…F. mortiferum 25557 was grown anaerobically (Gas Pak; BBL Microbiology Systems) at 37ЊC in modified Todd-Hewitt broth (34) supplemented with 0.25% (wt/vol) of the desired carbohydrate. Prereduced medium was inoculated with 10% (vol/vol) culture, and cells were grown to stationary phase (ϳ22 h).…”

“…Recent studies of carbohydrate metabolism by pathogenic fusobacteria (33,34,45) resulted, fortuitously, in the first preparative isolation of maltose 6P from any bacterial species (32). However, our attempts to demonstrate the enzymatic cleavage of the phosphorylated disaccharide were initially unsuccessful.…”

Metabolism of sucrose and its five isomers by Fusobacterium mortiferum

“…Fusobacteria are Gram-negative anaerobic rods that are usually described as weakly or asaccharolytic, and most species, including Fusobacterium nucleatum, use amino acids as fermentable energy sources (Robrish et al, 1987 ;Robrish & Thompson, 1990). The products of metabolism (acetic, butyric and propionic acids) may penetrate periodontal tissue, thereby contributing to the aetiology of gingivitis and periodontal disease.…”