Thermostable enzymes offer potential benefits in the hydrolysis of lignocellulosic substrates; higher specific activity decreasing the amount of enzymes, enhanced stability allowing improved hydrolysis performance and increased flexibility with respect to process configurations, all leading to improvement of the overall economy of the process. New thermostable cellulase mixtures were composed of cloned fungal enzymes for hydrolysis experiments. Three thermostable cellulases, identified as the most promising enzymes in their categories (cellobiohydrolase, endoglucanase and beta-glucosidase), were cloned and produced in Trichoderma reesei and mixed to compose a novel mixture of thermostable cellulases. Thermostable xylanase was added to enzyme preparations used on substrates containing residual hemicellulose. The new optimised thermostable enzyme mixtures were evaluated in high temperature hydrolysis experiments on technical steam pretreated raw materials: spruce and corn stover. The hydrolysis temperature could be increased by about 10-15 degrees C, as compared with present commercial Trichoderma enzymes. The same degree of hydrolysis, about 90% of theoretical, measured as individual sugars, could be obtained with the thermostable enzymes at 60 degrees C as with the commercial enzymes at 45 degrees C. Clearly more efficient hydrolysis per assayed FPU unit or per amount of cellobiohydrolase I protein used was obtained. The maximum FPU activity of the novel enzyme mixture was about 25% higher at the optimum temperature at 65 degrees C, as compared with the highest activity of the commercial reference enzyme at 60 degrees C. The results provide a promising basis to produce and formulate improved enzyme products. These products can have high temperature stability in process conditions in the range of 55-60 degrees C (with present industrial products at 45-50 degrees C) and clearly improved specific activity, essentially decreasing the protein dosage required for an efficient hydrolysis of lignocellulosic substrates. New types of process configurations based on thermostable enzymes are discussed.