S112: The quest for hyperthermophilic cellulases

 Samples were collected from a continental geothermal source in Nevada, USA, at 94ºC. Primary enrichment was carried out at 90ºC in mineral media with crystalline cellulose as sole carbon source and secondary enrichments utilizing CMC, Avicel, and filter paper as sole carbon sources at 90ºC.  The J1 consortium with three hyperthermophilic crenarchaeota capable of deconstructing crystalline cellulose at 94° C was grown in 20L batch.   A 90 kDa, multi-domain cellulase annotated as a member of the TIM barrel glycosyl hydrolase superfamily, was cloned and characterized following metagenomic studies on the consortium.  Two of the four domains of the enzyme had no homologs in the non-redundant database and the multi-domain architecture of this protein is uncommon for hyperthermophilic endoglucanases.  The recombinant enzyme had optimal activity at 109ºC, a half-life of 5 hours at 100ºC, and remained active in the presence of strong detergents, high salt concentrations, and ionic liquids.  Illumina sequencing of DNA from the enrichment revealed that a consortium of three Archaea was present. The genome of the dominant organism, Candidatus Synarcheum cellulolyticum,  was closed, and consisted of a 1.89 Mb chromosome and a 21 kb plasmid. The chromosome encodes an unusual multi-domain cellulase with a optimum temperature of 109° C as well an extensive set of glycosyl hydrolases, which are predicted to deconstruct and/or hydrolyze cellulose or hemicellulose.The hyperthermophiles represent the first instance of Archaea that are able to hydrolyze and grow on crystalline cellulose optimally above 90ºC.