|Chen, J - UNIV WISCONIN, MADISON|
|Stevenson, D - UNIV WISCONSIN, MADISON|
Submitted to: Conference on Rumen Function
Publication Type: Abstract Only
Publication Acceptance Date: November 15, 2000
Publication Date: N/A
Technical Abstract: Inhibition of growth of R.flavefaciens strains FD-1 and B34b by R. albus 7 is due to a heat-labile, protease sensitive agent, suggesting that the inhibitor is a bacteriocin. The agent was purified from culture supernatants of R. albus 7 by ammonium sulfate precipitation, ultrafiltration, gel filtration, and anion-exchange chromatography. The active fraction from the last step contained two proteins (MW ~36kDa and ~45kDa). The 36kda protein contained the inhibitory activity, and was purified by electroblotting and subjected to N-terminal and internal sequencing. BLAST sequence analysis revealed no homology to small, cationic bacteriocins, but weak homology to some clostridium and Ruminococcus endoglucanases. Oligonucleotide probes, designed from the N-terminal amino acid sequence, hybridized to a 4 kb HindIII fragment of R. albus 7 DNA. Fragments in this size range were ligated into pGEM-3Z and electrotransformed into E. coli JM109. Transformants were screened by colony hybridization, and one was found to contain a cloned ~1.4 kb HindIII fragment that may contain the gene for the bacteriocin.