Lucigen and Great Lakes Bioenergy Research Center

Middleton, WI, United States

Lucigen and Great Lakes Bioenergy Research Center

Middleton, WI, United States
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Brumm P.,C5 6 Technologies and Great Lakes Bioenergy Research Center | Hermanson S.,C5 6 Technologies and Great Lakes Bioenergy Research Center | Hochstein B.,Montana State University | Boyum J.,Lucigen and Great Lakes Bioenergy Research Center | And 3 more authors.
Applied Biochemistry and Biotechnology | Year: 2011

The genome of Dictyoglomus turgidum was sequenced and analyzed for carbohydrases. The broad range of carbohydrate substrate utilization is reflected in the high number of glycosyl hydrolases, 54, and the high percentage of CAZymes present in the genome, 3.09% of its total genes. Screening a random clone library generated from D. turgidum resulted in the discovery of five novel biomass-degrading enzymes with low homology to known molecules. Whole genome sequencing of the organism followed by bioinformatics-directed amplification of selected genes resulted in the recovery of seven additional novel enzyme molecules. Based on the analysis of the genome, D. turgidum does not appear to degrade cellulose using either conventional soluble enzymes or a cellulosomal degradation system. The types and quantities of glycosyl hydrolases and carbohydrate-binding modules present in the genome suggest that D. turgidum degrades cellulose via a mechanism similar to that used by Cytophaga hutchinsonii and Fibrobacter succinogenes. © 2010 Springer Science+Business Media, LLC.


Mead D.,Lucigen and Great Lakes Bioenergy Research Center | Boyum J.,Lucigen and Great Lakes Bioenergy Research Center | Drinkwater C.,Lucigen and Great Lakes Bioenergy Research Center | Gowda K.,Lucigen and Great Lakes Bioenergy Research Center | And 2 more authors.
Applied Biochemistry and Biotechnology | Year: 2011

Fibrobacter succinogenes is a cellulolytic bacterium that degrades plant cell wall biomass in ruminant animals and is among the most rapidly fibrolytic of all mesophilic bacteria. The complete genome sequence of Fisuc was completed by the DOE Joint Genome Institute in late 2009. Using new expression tools developed at Lucigen and C5-6 Technologies and a multi-substrate screen, 5,760 random shotgun expression clones were screened for biomass-degrading enzymes, representing 2x genome expression coverage. From the screen, 169 positive hits were recorded and 33 were unambiguously identified by sequence analysis of the inserts as belonging to CAZy family genes. Eliminating duplicates, 24 unique CAZy genes were found by functional screening. Several previously uncharacterized enzymes were discovered using this approach and a number of potentially mis-annotated enzymes were functionally characterized. To complement this approach, a high-throughput system was developed to clone and express all the annotated glycosyl hydrolases and carbohydrate esterases in the genome. Using this method, six previously described and five novel CAZy enzymes were cloned, expressed, and purified in milligram quantities. © 2010 Springer Science+Business Media, LLC.

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