6-03: Mining Clostridium thermocellum for Enzymatically Active Carbohydrases

The efficient hydrolysis of biomass to 5 carbon and 6 carbon sugars is limited by the lack of affordable, high specific activity enzymes. Screening of genomic and metagenoic libraries for new biomass-degrading enzymes has had only limited success.  We examined a number of screening strategies using Clostridium thermocellum (Cth) as a target-rich model organism to validate the efficiency of capturing carbohydrases that may prove useful for biomass degradation. The Cth genome has been sequenced and is predicted to have genes for over 60 potential biomass-degrading enzymes associated with the cellulosome, and another 18 enzymes that are noncellulosomal. Two different cloning systems were used for gene expression and two different screening methods were utilized for identification of positive clones. A comparison of the methods showed large differences both in the total number of positive clones identified as well as large differences in the total number of different enzymes captured.  This poster also describes the gene products that were captured by the individual screens.