Novel Penicillium cellulases for total hydrolysis of lignocellulosics

• Two novel Penicillium strains (TUB F-2220 and TUB F-2378) with high cellulase production were discovered.
• Enzyme mixtures of both Penicillium strains hydrolyzed efficiently of crystalline cellulose, were similar in hydrolysis of wheat straw but differed in hydrolysis of spruce.
• Spruce lignin inhibited especially the F-2378 enzyme mixture in temperature dependent manner.
• The main proteins in the TUB F-2220 (Penicillium pulvillorum) cultures were GH7 and GH6 CBHs (PpCBH1 and PpCBH2).
• Adsorption of PpCBH1 on spruce lignin was pH and temperature dependent.

The (hemi)cellulolytic systems of two novel lignocellulolytic Penicillium strains (Penicillium pulvillorum TUB F-2220 and P. cf. simplicissimum TUB F-2378) have been studied. The cultures of the Penicillium strains were characterized by high cellulase and β-glucosidase as well moderate xylanase activities compared to the Trichoderma reesei reference strains QM 6a and RUTC30 (volumetric or per secreted protein, respectively). Comparison of the novel Penicillium and T. reesei secreted enzyme mixtures in the hydrolysis of (ligno)cellulose substrates showed that the F-2220 enzyme mixture gave higher yields in the hydrolysis of crystalline cellulose (Avicel) and similar yields in hydrolysis of pre-treated spruce and wheat straw than enzyme mixture secreted by the T. reesei reference strain. The sensitivity of the Penicillium cellulase complexes to softwood (spruce) and grass (wheat straw) lignins was lignin and temperature dependent: inhibition of cellulose hydrolysis in the presence of wheat straw lignin was minor at 35 °C while at 45 °C by spruce lignin a clear inhibition was observed. The two main proteins in the F-2220 (hemi)cellulase complex were partially purified and identified by peptide sequence similarity as glycosyl hydrolases (cellobiohydrolases) of families 7 and 6. Adsorption of the GH7 enzyme PpCBH1 on cellulose and lignins was studied showing that the lignin adsorption of the enzyme is temperature and pH dependent. The ppcbh1 coding sequence was obtained using PCR cloning and the translated amino acid sequence of PpCBH1 showed up to 82% amino acid sequence identity to known Penicillium cellobiohydrolases.