Two novel GH11 endo-xylanases from Myceliophthora thermophila C1 act differently toward soluble and insoluble xylans

Two novel GH11 endo-xylanases from Myceliophthora thermophila C1 (C1), Xyl7 and Xyl8, were purified and the influence of solubility and molecular structure of various xylans on their efficiency was investigated. Both endo-xylanases were hindered by a high degree of substitution of a xylan. The two GH11 xylanases released different products from the xylans, in which Xyl7 displayed a degradation product composition closer to GH10 xylanases. A correlation of the degradation product composition with a specific residue at position 163 in the amino acid sequence of Xyl8 is suggested: tyrosine in Xyl8; valine in Xyl7. This is confirmed with examples of various endo-xylanases reported in literature. The C1 GH11 xylanases were more efficient on self-associated xylan compared to C1 GH10 endo-xylanases and they released more small xylooligomers from these xylans. This is contrary to the general assumption that GH10 xylanases degrade xylans to a higher degree than GH11 xylanase

Two GH10 endo-xylanases from Myceliophthora thermophila C1 with and without cellulose binding module act differently towards soluble and insoluble xylans

Xylanases are mostly classified as belonging to glycoside hydrolase (GH) family 10 and 11, which differ in catalytic properties and structures. However, within one family, differences may also be present. The influence of solubility and molecular structure of substrates towards the efficiency of two GH10 xylanases from Myceliophthora thermophila C1 was investigated. The xylanases differed in degradation of high and low substituted substrate and the substitution pattern was an important factor influencing their efficiency. Alkali-labile interactions, as well as the presence of cellulose within the complex cell wall structure hindered efficient hydrolysis for both xylanases. The presence of a carbohydrate binding module did not enhance the degradation of the substrates. The differences in degradation could be related to the protein structure of the two xylanases. The study shows that the classification of enzymes does not predict their performance towards various substrates

Two novel GH11 endo-xylanases from Myceliophthora thermophila C1 act differently toward soluble and insoluble xylans

Two novel GH11 endo-xylanases from Myceliophthora thermophila C1 (C1), Xyl7 and Xyl8, were purified and the influence of solubility and molecular structure of various xylans on their efficiency was investigated. Both endo-xylanases were hindered by a high degree of substitution of a xylan. The two GH11 xylanases released different products from the xylans, in which Xyl7 displayed a degradation product composition closer to GH10 xylanases. A correlation of the degradation product composition with a specific residue at position 163 in the amino acid sequence of Xyl8 is suggested: tyrosine in Xyl8; valine in Xyl7. This is confirmed with examples of various endo-xylanases reported in literature. The C1 GH11 xylanases were more efficient on self-associated xylan compared to C1 GH10 endo-xylanases and they released more small xylooligomers from these xylans. This is contrary to the general assumption that GH10 xylanases degrade xylans to a higher degree than GH11 xylanase

Two GH10 endo-xylanases from Myceliophthora thermophila C1 with and without cellulose binding module act differently towards soluble and insoluble xylans

Xylanases are mostly classified as belonging to glycoside hydrolase (GH) family 10 and 11, which differ in catalytic properties and structures. However, within one family, differences may also be present. The influence of solubility and molecular structure of substrates towards the efficiency of two GH10 xylanases from Myceliophthora thermophila C1 was investigated. The xylanases differed in degradation of high and low substituted substrate and the substitution pattern was an important factor influencing their efficiency. Alkali-labile interactions, as well as the presence of cellulose within the complex cell wall structure hindered efficient hydrolysis for both xylanases. The presence of a carbohydrate binding module did not enhance the degradation of the substrates. The differences in degradation could be related to the protein structure of the two xylanases. The study shows that the classification of enzymes does not predict their performance towards various substrates

Purification, cloning and characterisation of two forms of thermostable and highly active cellobiohydrolase I (Cel7A) produced by the industrial strain of Chrysosporium lucknowense

Two forms of cellobiohydrolase I (CBH I, Cel7A) were purified from the culture ultrafiltrate of a mutant strain of the fungus Chrysosporium lucknowense, an industrial producer of cellulases and hemicellulases. The enzymes had different molecular masses (52 and 65 kDa, SDS-PAGE data) but the same pI (4.5). Peptide sequencing showed that a single gene encodes both proteins. Both enzymes displayed maximum activity at pH 5.0-5.5; they had similar specific activities against soluble substrates. However, the 65 kDa CBH I was much more efficient in hydrolysis of Avicel and cotton cellulose, and its adsorption ability on Avicel was notably higher in comparison to the 52 kDa enzyme. Using the in-gel tryptic digestion followed by MALDI-TOF mass-spectrometry, it was shown that the 52 kDa enzyme represents the core catalytic module of the intact 65 kDa CBH I without a cellulose-binding module and major part of glycosylated linker. Both enzymes were stable at 50°C for 24 h. At higher temperature, the 65 kDa enzyme showed better thermostability: it retained >90% of activity after 7 h at 60°C and 50% of activity after 3 h at 65°C. The intact CBH I is also notably more thermostable than the Trichoderma reesei CBH I (by 6°C, the data of differential scanning microcalorimetry study). The cbh1 gene was cloned and then the amino acid sequence of Cel7A was deduced from the gene sequence. The enzyme had high degree of similarity (up to 74%) to family 7 cellobiohydrolases and lower degree of similarity (up to 41%) to family 7 endoglucanases. © 2004 Elsevier Inc. All rights reserved