Tuesday, May 3, 2011
Nitrogen and sulfur are required to maintain microbial biocatalysts for consolidated bioprocessing of lignocellulose. After anaerobic fermentation and product extraction, both elements will be enriched in the residue, and subsequent combustion will produce NOx and SO2 gases. These gas emissions are regulated, making it desirable to sequester and deplete their precursors during fermentation. Defined minimal growth media were developed for batch cultures of Clostridium thermocellum and Caldicellulosiruptor bescii, two thermophilic cellulose-degrading bacteria. Nitrogen and sulfur requirements were identified for each strain: both organisms used sulfate as a sole sulfur source through assimilatory sulfate reduction. Minimal vitamin and cofactor requirements were identified by metabolic reconstruction from genome sequences and validated experimentally. The new defined growth medium for C. thermocellum efficiently supported ethanolic fermentation using crystalline cellulose and acid-pretreated poplar. Iron limitation reduced the production of yellow affinity substance by C. thermocellum grown on crystalline cellulose. The defined minimal media facilitated cocultures of C. thermocellum or C. bescii with compatible thermophilic bacteria and simplified fermentation product analysis. Polyclonal antibodies and activity assays were used to identify and quantify C. bescii cells in cocultures with other cellulolytic and proteolytic bacteria. These results support developing microbial cocultures to enhance bioprocessing, improve fermentation product yields, provide new means of genetically engineering systems, and reduce fermentation and downstream waste processing costs.