Utilization of Chondroitin Sulfate byBacteroides thetaiotaomicronGrowing in Carbohydrate-Limited Continuous Culture

Abstract

When Bacteroides thetaiotaomicron, an obligate anaerobe from the human colonic flora, was grown in continuous culture with the mucopolysaccharide chondroitin sulfate as the limiting source of carbohydrate, growth yields ranged from 48 g of cell dry weight per mol of equivalent monosaccharide at a growth rate of 3.5 h per generation to 32 g per mol at a growth rate of 24 h per generation. The theoretical maximum growth yield (61 g of cell dry weight per mol of equivalent monosaccharide) was comparable to that of 54 g per mol, which was obtained previously when glucuronic acid, a component of chondroitin sulfate, was the limiting carbohydrate (S. F. Kotarski and A. A. Salyers, J. Bacteriol. 146:853-860, 1981). However, the maintenance coefficient was three times higher when chondroitin sulfate was the substrate than when glucuronic acid was the substrate. The specific activity of chondroitin lyase (EC 4.2.2.4), an enzyme which cleaves chondroitin sulfate into disaccharides, declined by nearly 50%o as growth rates decreased from 3.5 to 24 h per generation. By contrast, the specific activities of several glycolytic enzymes and disaccharidases remained constant over this range of growth rates. Although chondroitin sulfate was growth limiting, some carbohydrate was detectable in the extracellular fluid at all growth rates. At rapid growth rates (1 to 2 h per generation), this residual carbohydrate included fragments of chondroitin sulfate having a wide range of molecular weights. At slower growth rates (2 to 24 h per generation), the residual carbohydrate consisted mainly of a small fragnent which migrated on paper chromatograms more slowly than the disaccharides produced by chondroitin lyase but faster than a tetrasaccharide. This small fragment may represent the reducing end of the chondroitin sulfate molecule. 1008 on July 16, 2020 by guest

Digestion of proteoglycan by Bacteroides thetaiotaomicron

“…Paper chromatography of this low-molecular-weight material revealed that it did not consist of unsaturated disaccharides (i.e., products of digestion of chondroitin sulfate by chondroitin lyase) but consisted of a compound which ran more slowly than ADi-4S on the paper chromatogram. This oligomer comigrated with an oligomer which we found previously in the extracellular fluid from bacteria which were grown in continuous culture with chondroitin sulfate as the sole carbohydrate source (16).…”

“…To determine whether chondroitin lyase was being released from the cells during incubation, samples of the extracellular fluid were tested for lyase activity by incubating 0.2-mi portions with chondroitin sulfate A and monitoring the increase in absorbance at 235 nm (16). To check for cell lysis during incubation, extracellular fluid was also assayed to detect the cytoplasmic enzyme phosphoglucose isomerase (11).…”

“…This cell-associated catabolic system appears to work well when isolated, linear polymers are being degraded. When B. thetaiotaomicron is grown on chondroitin sulfate, cell yields and growth rates are comparable to those attained when the bacteria are grown on monosaccharide components of chondroitin (7,16). In nature, however, chondroitin sulfate molecules are bound up in com-plex aggregates called proteoglycans (8,13).…”

“…Bacteroides thetaiotaomicron, a gram-negative anaerobe which is found in human colons, can utilize mucopolysaccharides, such as chondroitin sulfate, as sole sources of carbon and energy, and it has been suggested that mucopolysaccharides which are released due to turnover of the intestinal mucosa of the host may be a natural carbon source for these organisms (16,17). Previous work has shown that all of the enzymes which are involved in the catabolism of chondroitin sulfate by B. thetaiotaomicron are cell associated rather than extracellular.…”

Distinct polysaccharide growth profiles of human intestinalPrevotella copriisolates

“…Although beta-glucanases break down linkages in beta-glucan, and alpha-L-arabinofuranosidases break down linkages in rhamnogalacturonan and arabinan, a polysaccharide substrate containing both linkages has not been described. In contrast with several Bacteroides species, which catabolize polysaccharides from both animal and plant sources, all identified pPUL substrates for P. copri isolates were from plant sources, and none were from animal sources (Martens et al, 2011;Salyers AA, 1982).…”