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oai:www.ideals.illinois.edu:2142/21603 2023-07-10 col_2142_5131 col_2142_14771 com_2142_5130 com_2142_685 com_2142_397

Restriction data tranferred 2014-07-01T11:23:52-05:00 Original Data Group with Access UIUC Users [automated] Release Date: none Reason: ETDs are only available to UIUC Users without author permission ETDs are only available to UIUC Users without author permission U of I Only AAI9215803 (UMI)AAI9215803 http://hdl.handle.net/2142/21603 eng Copyright 1992 Doerner, Kinchel Clay Biology, Microbiology Mackie, Roderick I. Doerner, Kinchel Clay 2011-05-07T13:13:35Z 2011-05-07T13:13:35Z 10000-01-01 1992 The cellulase system of the anaerobic bacterium Ruminococcus flavefaciens FD-1 was investigated. Through the use of ion-exchange chromatography and non-denaturing polyacrylamide gel electrophoresis, crude cellulase was found to contain no fewer than 18 endo-$\beta$-1,4-glucanase components and present in two cellulase complexes. No synergistic effect on $\sp{14}$C-cellulose degradation was observed when these two cellulase complexes were incubated together or when the cellulase complexes were incubated together or separately in the presence of an exo-$\beta$-1,4-glucanase produced by this organism (Exoglucanase A). Both cellulase complexes degraded xylan and the xylanase components migrated coincidently with all endo-$\beta$-1,4-glucanase components present in both cellulase complexes. A monoclonal antibody (MAb S1) generated against Exoglucanase A was used to investigate cellulolysis by R. flavefaciens. MAb S1 recognized few components in crude cellulase demonstrating its specificity for Exoglucanase A. MAb S1 was used to investigate the cellular location of the Exoglucanase A. Thin-sectioning of R. flavefaciens followed by immunolabelling, and examination using a transmission electron microscope showed a non-specific interaction of the monoclonal antibody for the embedding resin. Although, various treatments were tried to prevent this interaction, no conclusions about the cellular location of Exoglucanase A could be drawn. MAb S1 also inhibited the action of Exoglucanase A in vitro but failed to inhibit growth of R. flavefaciens when added to culture medium. This suggests the action of Exoglucanase A is not necessary for growth or the antibody concentration in the medium was too low to affect growth. Nutritional factors affecting cellulase expression in R. flavefaciens were also investigated. Cellobiose-grown cells produced more endo-$\beta$-1,4-glucanase and $\sp{14}$C-cellulase, but an equal amount of exo-$\beta$-1,4-glucanase compared to cellulose-grown cells. Growth on cellobiose or cellulose did not affect cellular location of either endo-$\beta$-1,4-glucanase or exo-$\beta$-1,4-glucanase, however cellulose-grown cells exhibited higher $\sp{14}$C-cellulase levels present in the supernatant as opposed to the cell pellet. Furthermore, addition of cellulose to a growing culture using cellobiose did not increase cellulase production compared to a control with no addition. Cellulase was constitutively produced in cellobiose- and cellulose-grown R. flavefaciens cells. Made available in DSpace on 2011-05-07T13:13:35Z (GMT). No. of bitstreams: 2 license.txt: 4922 bytes, checksum: 910b249b4beec47e7ab768910c8f966f (MD5) 9215803.pdf: 8710012 bytes, checksum: 8e6cfc5fbae20dcc2c33589d54ae74f3 (MD5) Previous issue date: 1992 Item marked as restricted to the 'UIUC Users [automated]' Group (id=2) by Howard Ding (hding2@illinois.edu) on 2011-05-07T14:51:54Z Item is restricted indefinitely. Agriculture, Animal Culture and Nutrition Investigation of Ruminococcus flavefaciens FD-1 cellulase text Animal Sciences Animal Sciences University of Illinois at Urbana-Champaign Dissertation Ph.D.