Insights into the action of phylogenetically diverse microbial expansins on the structure of cellulose microfibrils.

Background: Microbial expansins (EXLXs) are non-lytic proteins homologous to plant expansins involved in plant cell wall formation. Due to their non-lytic cell wall loosening properties and potential to disaggregate cellulosic structures, there is considerable interest in exploring the ability of microbial expansins (EXLX) to assist the processing of cellulosic biomass for broader biotechnological applications. Herein, EXLXs with different modular structure and from diverse phylogenetic origin were compared in terms of ability to bind cellulosic, xylosic, and chitinous substrates, to structurally modify cellulosic fibrils, and to boost enzymatic deconstruction of hardwood pulp.

Fungal loosenin-like proteins boost the cellulolytic enzyme conversion of pretreated wood fiber and cellulosic pulps.

Microbial expansin-related proteins, including loosenins, can disrupt cellulose networks and increase enzyme accessibility to cellulosic substrates. Herein, four loosenins from Phanerochaete carnosa (PcaLOOLs), and a PcaLOOL fused to a family 63 carbohydrate-binding module, were compared for ability to boost the cellulolytic deconstruction of steam pretreated softwood (SSW) and kraft pulps from softwood (ND-BSKP) and hardwood (ND-BHKP). Amending the Cellic® CTec-2 cellulase cocktail with PcaLOOLs increased reducing products from SSW by up to 40 %, corresponding to 28 % higher glucose yield.

Enzyme kinetics by GH7 cellobiohydrolases on chromogenic substrates is dictated by non-productive binding: insights from crystal structures and MD simulation.

Cellobiohydrolases (CBHs) in the glycoside hydrolase family 7 (GH7) (EC3.2.1.

Discovery of fungal oligosaccharide-oxidising flavo-enzymes with previously unknown substrates, redox-activity profiles and interplay with LPMOs.

Oxidative plant cell-wall processing enzymes are of great importance in biology and biotechnology. Yet, our insight into the functional interplay amongst such oxidative enzymes remains limited. Here, a phylogenetic analysis of the auxiliary activity 7 family (AA7), currently harbouring oligosaccharide flavo-oxidases, reveals a striking abundance of AA7-genes in phytopathogenic fungi and Oomycetes.

Loss of AA13 LPMOs impairs degradation of resistant starch and reduces the growth of .

Background: Lytic polysaccharide monooxygenases (LPMOs) are often studied in simple models involving activity measurements of a single LPMO or a blend thereof with hydrolytic enzymes towards an insoluble substrate. However, the contribution of LPMOs to polysaccharide breakdown in complex cocktails of hydrolytic and oxidative enzymes, similar to fungal secretomes, remains elusive. Typically, two starch-specific AA13 LPMOs are encoded by mainly Ascomycota genomes.

A novel starch-binding laccase from the wheat pathogen Zymoseptoria tritici highlights the functional diversity of ascomycete laccases.

Background: Laccases are multicopper oxidases, which are assigned into auxiliary activity family 1 (AA1) in the CAZy database. These enzymes, catalyzing the oxidation of phenolic and nonphenolic substrates coupled to reduction of O to HO, are increasingly attractive as eco-friendly oxidation biocatalysts. Basidiomycota laccases are well characterized due to their potential in de-lignification of lignocellulose.

Biochemical and Structural Characterizations of Two Dictyostelium Cellobiohydrolases from the Amoebozoa Kingdom Reveal a High Level of Conservation between Distant Phylogenetic Trees of Life.

Unlabelled: Glycoside hydrolase family 7 (GH7) cellobiohydrolases (CBHs) are enzymes commonly employed in plant cell wall degradation across eukaryotic kingdoms of life, as they provide significant hydrolytic potential in cellulose turnover. To date, many fungal GH7 CBHs have been examined, yet many questions regarding structure-activity relationships in these important natural and commercial enzymes remain. Here, we present the crystal structures and a biochemical analysis of two GH7 CBHs from social amoeba: Dictyostelium discoideum Cel7A (DdiCel7A) and Dictyostelium purpureum Cel7A (DpuCel7A).

Structural insights into the inhibition of cellobiohydrolase Cel7A by xylo-oligosaccharides.

Unlabelled: The filamentous fungus Hypocrea jecorina (anamorph of Trichoderma reesei) is the predominant source of enzymes for industrial saccharification of lignocellulose biomass. The major enzyme, cellobiohydrolase Cel7A, constitutes nearly half of the total protein in the secretome. The performance of such enzymes is susceptible to inhibition by compounds liberated by physico-chemical pre-treatment if the biomass is kept unwashed.

Expression, crystal structure and cellulase activity of the thermostable cellobiohydrolase Cel7A from the fungus Humicola grisea var. thermoidea.

Glycoside hydrolase family 7 (GH7) cellobiohydrolases (CBHs) play a key role in biomass recycling in nature. They are typically the most abundant enzymes expressed by potent cellulolytic fungi, and are also responsible for the majority of hydrolytic potential in enzyme cocktails for industrial processing of plant biomass. The thermostability of the enzyme is an important parameter for industrial utilization.

The mechanism of cellulose hydrolysis by a two-step, retaining cellobiohydrolase elucidated by structural and transition path sampling studies.

Glycoside hydrolases (GHs) cleave glycosidic linkages in carbohydrates, typically via inverting or retaining mechanisms, the latter of which proceeds via a two-step mechanism that includes formation of a glycosyl-enzyme intermediate. We present two new structures of the catalytic domain of Hypocrea jecorina GH Family 7 cellobiohydrolase Cel7A, namely a Michaelis complex with a full cellononaose ligand and a glycosyl-enzyme intermediate, that reveal details of the 'static' reaction coordinate. We also employ transition path sampling to determine the 'dynamic' reaction coordinate for the catalytic cycle.

Adaptation of Dekkera bruxellensis to lignocellulose-based substrate.

Adaptation of Dekkera bruxellensis to lignocellulose hydrolysate was investigated. Cells of D. bruxellensis were grown for 72 and 192 H in batch and continuous culture, respectively (adapted cells).

Structural, biochemical, and computational characterization of the glycoside hydrolase family 7 cellobiohydrolase of the tree-killing fungus Heterobasidion irregulare.

Root rot fungi of the Heterobasidion annosum complex are the most damaging pathogens in temperate forests, and the recently sequenced Heterobasidion irregulare genome revealed over 280 carbohydrate-active enzymes. Here, H. irregulare was grown on biomass, and the most abundant protein in the culture filtrate was identified as the only family 7 glycoside hydrolase in the genome, which consists of a single catalytic domain, lacking a linker and carbohydrate-binding module.

Improved bio-energy yields via sequential ethanol fermentation and biogas digestion of steam exploded oat straw.

Using standard laboratory equipment, thermochemically pretreated oat straw was enzymatically saccharified and fermented to ethanol, and after removal of ethanol the remaining material was subjected to biogas digestion. A detailed mass balance calculation shows that, for steam explosion pretreatment, this combined ethanol fermentation and biogas digestion converts 85-87% of the higher heating value (HHV) of holocellulose (cellulose and hemicellulose) in the oat straw into biofuel energy. The energy (HHV) yield of the produced ethanol and methane was 9.