Studying Anaerobic Fungi for use in Biofuel Applications

Anaerobic fungi found in the guts of herbivorous mammals provide perhaps the most promising mechanism to break down plant biomass for conversion to biofuels and other chemical commodities. However, very little is known about these organisms and the details of their molecular structure. This research focuses on the characterization of anaerobic gut fungi and how they mediate the breakdown of cellulose, the predominate component of plant biomass, into sugars. While many of the enzymes that provide the cellulolytic capability of these organisms have been identified, there are few details on how these enzymes assemble to form large complex structures, and this is thought to contribute significantly to plant biomass break down. In anaerobic bacteria, cellulases assemble by non-catalytic domains called dockerins. Dockerins bind to cohesin domains encoded on a membrane-anchored scaffoldin.  In anaerobic fungi, dockerins have been identified, but the dockerin-binding cohesin and scaffoldin molecules have remained elusive, and this has been a major lack of understanding in these systems. To identify dockerin-binding cohesin molecules, we used a chemical crosslinker to trap protein-protein interactions between a purified dockerin and its binding partners. This method revealed a putative cohesin molecule with a molecular weight of approximately 75 kDa. The identity of this cohesin is currently being obtained by mass spectrometry. This new information on the molecular assembly of cellulose-degrading complexes may be exploited for use in the conversion of agricultural waste into biofuel and other value-added chemicals.