Biomass Depolymerization projects

Analysis of Bovine Rumen Microbiota Under Different Dietary Regimens for Identification of Feedstock-Targeted Cellulolytic Genes - Completed

In the gut environments of ruminants, conversion of cellulosic biomass into fermentable sugars takes place rapidly and at moderate pH and temperature. This team centered at the DOE Joint Genome Institute identified those enzymes produced by the abundant microbes in ruminant guts; enzymes that can potentially be co-opted for cellulosic biomass conversion. In this study, switchgrass and control substrates in synthetic porous bags were incubated in the bovine rumen and feedstuff-associated microbes were sampled for DNA and RNA extraction.

project Highlights

2011 Highlights

Rubin and his postdoctoral fellow, Matthias Hess, collaborated with Berkeley professor Doug Clark’s group to characterize the carbohydrate-active enzymes found in the rumen of cows fed on switchgrass. After sequencing approximately 260 billon base pairs of DNA, they concluded that at least 1,000 organisms were present in the rumen samples and that they were producing at least 27,500 carbohydrate-active enzymes that had not been previously described.

2010 Highlights

Rubin’s group sequenced and analyzed more than a quarter terabase of metagenomic DNA generated directly from microbes adherent to plant fiber in the cow rumen. They identified 27,755 candidate genes encoding putative carbohydrate-active enzymes. They synthesized 90 candidate genes selected from this pool and tested the activity of the expressed proteins against 10 substrates, including the potential biofuel feedstocks miscanthus and switchgrass. The authenticity of identified novel genes and individual assembled draft genomes was validated by complementary methods including single-cell genome sequencing.

2009 Highlights

After incubating synthetic nylon bags of switchgrass in the rumen of a fistulated cow for three days, Rubin’s team studied the fiber-adherent microbial community and the genes encoded in the microbial genomes. Rubin’s group observed that cellulose, hemicellulose, and lignin components of switchgrass are degraded within the bovine rumen, supporting the hypothesis that the rumen microbiome as a valuable source for lignocellulolytic proteins. They successfully recovered the full-length gene sequence of 119 endoglucanases, 108 beta-glucosidases, and 8 cellobiohydrolases. The cellulolytic enzymes identified from the switchgrass-associated microbiome have a relatively low overall amino acid sequence identity (52%) to known lignocellulolytic enzymes, suggesting that these biocatalysts might have significantly different characteristics from known lignocellulases.

 

Publications

Published in 2011

Metagenomic Discovery of Biomass-Degrading Genes and Genomes from Cow Rumen, Matthias Hess, Alexander Sczyrba, Rob Egan, Tae-Wan Kim, Harshal Chokhawala, Gary Schroth, Shujun Luo, Douglas S. Clark, Feng Chen, Tao Zhang, Roderick Mackie, Len Pennacchio, Susannah Tringe, Axel Visel, Tanja Woyke, Zhong Wang, Edward M. Rubin, Science, 331: (6016), pps. 463-467, January 28, 2011.

Published in 2010

Functional Diversity of Four Glycoside Hydrolase Family 3 Enzymes from the Rumen Bacterium Prevotella bryantii B14, Dylan Dodd, Shinichi Kiyonari, Roderick I. Mackie, Isaac K. O. Cann, Journal of Bacteriology, 192(9): pp. 2335-2345, May 2010.

 


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