Functional genomics of the Clostridium thermocellum cellulosome

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The cellulosome of the thermophilic bacterium Clostridium thermocellum is a large, extracellular multienzyme complex consisting of more than 40 different modular enzymes, arranged on a non-catalytic structural protein (scaffoldin). The complex structure of the cellulosome is the basis for its extreme efficiency in the hydrolysis of crystalline cellulose, probably due to local synergism. Therefore the cellulosome is of the highest interest for the conversion of biomass. The genome of C. thermocellum has been sequenced, making functional genomics feasable. This project plans 1) to determine and to identify all components of the cellulosome with proteomic methods. 2) New components will be cloned with reverse genetics, sequenced and expressed for biochemical characterisation. 3) The composition of the whole complex will be investigated in vivo: changes in protein pattern after growth on different substrates point to the cellulosomal components necessary for the hydrolysis of crystalline cellulose. 4) Essential components will also be identified by their absence in induced non-cellulolytic mutants. 5) Analysis of the cellulosome-components bound to the bacterial cell wall gives an indication for the mechanism of attachment and assembly.