The cellular location of Prevotella ruminicola beta-1,4-D-endoglucanase and its occurrence in other strains of ruminal bacteria

Abstract

Prevotella ruminicola B 1 4, TC1-1, TF1-3, and TS1-5 all produced immunologically cross-reacting 88-and 82-kDa carboxymethyl cellulases (CMCases). P. ruminicola 23, 118B, 20-63, and 20-78 had much lower CMCase activities, and Western blots (immunoblots) showed no cross-reaction with the B 1 4 CMCase antiserum. Fibrobacter succinogenes S85 and Selenomonas ruminantium HD4 and D produced CMCases, but these enzymes were smaller and did not cross-react with the B 1 4 CMCase antiserum. The B 1 4 CMCase antiserum inhibited the B 1 4, TC1-1, TF1-3, and TS1-5 CMCase activities and agglutinated these cells, but it had no effect on the other strains or species. On the basis of these results, the B 1 4 CMCase is a strain-specific enzyme that is located on the outside surface of the cells. P. ruminicola B 1 4 cultures, grown on sucrose, did not have significant CMCase activity, but these cells could bind purified 88-and 82-kDa CMCase but not 40.5-kDa CMCase. Because the 40.5-kDa CMCase is a fully active, truncated form of the CMCase, it appears that the N-terminal domain of the 88-kDa B 1 4 CMCase anchors the CMCase to the cells. Cells grown on cellobiose produced at least 10-fold more CMCase than the sucrose-grown cells, and the cellobiose-grown cells could only bind 15% as much CMCase as sucrose-grown cells. Virtually all of the CMCase activity of exponentially growing cultures was cell associated, but CMCase activity was eventually detected in the culture supernatant. On the basis of the observation that the 88-kDa CMCase was gradually converted to the 82-kDa CMCase when cultures reached the stationary phase without a change in specific activity, it appears that the 82-kDa protein is probably a proteolytic degradation product of the 88-kDa CMCase.

Interaction of molasses and monensin in alfalfa hay- or corn silage-based diets on rumen fermentation, total tract digestibility, and milk production by Holstein cows

“…Although CMCase is produced by other bacteria that do not degrade polymeric cellulose, we assumed that CMCase reflected a cumulative response to cellulolysis. In support, Prevotella ruminicola increased CMCase considerably when grown on cellobiose than on sucrose (Gardner et al, 1995), and complete degradation of cellulose probably involves a consortium of primary cellulolytic and secondary noncellulolytic bacteria (Wells et al, 1995). Changes in ruminal NDF degradability were proportionately associated with particle-bound CMCase activity (Huhtanen and Khalili, 1992;Piwonka et al, 1994).…”

Phylogenetic analysis of fiber-associated rumen bacterial community and PCR detection of uncultured bacteria

“…1). Prevotella species are recognized as the major groups in the rumen [15], and some Prevotella strains from the rumen possess endoglucanase and xylanase activities [16,17] but do not degrade cell wall material extensively in pure culture [15]. The isolation of many Prevotella-like clones from ruminally incubated hay stems suggests indirect concern of these bacteria with ruminal ¢ber breakdown possibly as oligosaccharide and xylan fermenters.…”

Changes in the Solid-, Liquid-, and Epithelium-Associated Bacterial Communities in the Rumen of Hu Lambs in Response to Dietary Urea Supplementation

“…This result is consistent with findings regarding the rumen solid and liquid fractions of dairy cattle fed ryegrass or white clover (Bowen et al, 2018) and a TMR (forage:concentrate = 70:30, forage = prewilted grass and maize silage) (De Mulder et al, 2017) and the rumen contents and epithelium of dairy cattle fed a TMR (forage:concentrate = 55:45, forage = corn silage and corn stover) (Liu et al, 2016). Prevotella represents one of the most abundant genera in the rumen; this genus exhibits genetic and metabolic diversity (Bekele et al, 2010) and plays roles in carbohydrate utilization (Dehority, 1966;Cotta, 1992;Gardner et al, 1995;Kabel et al, 2011), nitrogen metabolism (Kim et al, 2017), and fiber degradation (Mayorga et al, 2016). The results of these study suggest the importance of Prevotella spp.…”