The cellular location of Prevotella ruminicola beta-1,4-D-endoglucanase and its occurrence in other strains of ruminal bacteria.
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 CMCase, 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.
Gardner
R G
RG
Section of Biochemistry, Molecular and Cellular Biology, Cornell University, Ithaca, New York 14853, USA.
Wells
J E
JE
Russell
J B
JB
Wilson
D B
DB
D000818:Animals; D002462:Cell Membrane / Q000201:enzymology; D002480:Cellulase / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D004795:Enzyme Stability; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D006089:Gram-Negative Anaerobic Bacteria / Q000201:enzymology; D007150:Immunohistochemistry; D008970:Molecular Weight; D018720:Prevotella / Q000201:enzymology*; D012417:Rumen / Q000382:microbiology*; D013045:Species Specificity
D006026:Glycoside Hydrolases; D002480:Cellulase; C019826:carboxymethylcellulase