AbstractEnzymatic hydrolysis is a key step in bioethanol production. Efficient hydrolysis requires a consortium of different enzymes that are able to hydrolyze cellulose and hemicellulose into fermentable sugars. Myceliopthora thermophila is a promising candidate for the production of thermophilic cellulolytic enzymes, the use of which could reduce the cost of ethanol production. The growth conditions of the fungus were optimized in order to achieve increased secretion of extracellular cellulases. Optimal conditions were found to be 7.0% w/v brewer’s spent grain as the carbon source and 0.4% w/v ammonium sulfate as the nitrogen source. The cellulases obtained were characterized for their optimum activity. The optimum temperature and pH for cellulase activity are 65 °C and pH 5.5, respectively. Studies on thermal inactivation of the crude extract showed that the cellulases of M. thermophila are stable for temperatures up to 60 °C. At this temperature the half-life was found to be as high as 27 h. Enzymatic hydrolysis of cellulose resulted in 31.4% hydrolysis yield at 60 °C after 24 h of incubation. Finally, the recalcitrance constant for cellulose and cellulose pretreated with ionic liquids was calculated to be 5.46 and 2.69, respectively.