8-58: Biochemical, genomic and proteomic evaluation of a representative mucorale, the oleaginous Mucor circinelloides, for its cellulolytic potential

Lipid production by oleaginous microorganisms is a promising route to biodiesel production. However, most of them have to grow on sugars and agro-industrial wastes, and cannot direct utilize lignocellulosic substrates as carbon sources. Here, for the first time, we investigated the potential of Mucor circinelloides to assimilate cellulose and convert it into lipids. Our data show that M. circinelloides grew on CMC (carboxymethyl cellulose) plates, and its extracellular crude enzymes generated both glucose and cellobiose from CMC, but released only a small amount of cellobiose from Avicel, a crystalline cellulose with higher recalcitrance, indicating that it produced some, but not a full set of essential cellulases. To further evaluate Mucor’s cellulolytic potential, a genome-wide search revealed that totally 256 glycoside hydrolase (GH) proteins were predicted, including a set of core secretory cellulases - endoglucanases and β-glucosidases – but lack of exoglucanases such as CBH1 (cellobiohydrolase 1). However, none of above core secretory cellulases was found by our secretomic analysis; instead, the secretomic analysis identified a set of accessory cellulases, polysaccharide deacetylase and function unknown proteins, with a majority of them being N-glycosylated. Our data support a two-pronged strategy to overcome the above genomic, translational and post-translational bottlenecks for boosting Mucor’s cellulolytic capacity, by improving the expression of genome-predicted major endogenous cellulases, and by expressing foreign cellulases. Such strategy was preliminarily validated by observation that pre-saccharified Avicel and pre-saccharified dilute-acid pretreated corn stover supported fungal growth and lipid production in Mucor, and should be applicable to other oleaginous but non-cellulolytic microorganisms.