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The adsorption of lignin to cellulase is the major obstacle in the sugar-platform conversion of lignocellulosic bioresources. In this study, the adsorption behavior of untreated and pretreated lignin samples from corn stover to cellulase was investigated, in particular the effects of lignin adsorption on the composition and spatial conformation of free enzymes were explored. The results showed that pretreatments decreased the hydrophobic groups contents of lignin, i.e., aromatic ring, ether and carbonyl, as well as the content of ionizable group, i.e., carboxyl, which reduced its hydrophobicity and negative charge density, thus weakening the adsorption ability of lignin to cellulase. The lignin samples mainly adsorbed the CBHII component of cellulase to inhibit the synergistic effect of free enzymes. Lignin adsorption altered the spatial position of tryptophan residues in free enzymes, exposing them to the protein surface. In addition, the secondary structure of free enzymes was altered, with a decrease in the alpha-helix content and an increase in the random coil content, thus loosening the spatial conformation of free enzymes. The change degree in the spatial structure of free enzymes correlated with the adsorption capacity of the lignin, i.e., lignin with low adsorption capacity caused the least damage to free enzyme, with NaOH pretreated lignin being the best. It appears that appropriate pretreatment and chemical modification of enzymes to resist lignin adsorption is a promising long-term pathway to overcome the lignin inhibition during sugar-platform conversion of lignocellulosic bioresources. [ABSTRACT FROM AUTHOR]

[Display omitted] • DES improved cellulase activity by mainly rising βG activity with 20–35% increase. • DES improved cellulase stability by compacting the spatial structure. • DES formed a protective layer to prevent unproductive adsorption of lignin on cellulase. • DES significantly increased the saccharification efficiency of corn stover and poplar. The introduction of specific choline chloride (ChCl)- and betaine (Bet)-based deep eutectic solvents (DESs) into the conventional acetic acid–sodium acetate solution led to a significant increase in cellulase activity and stability. In particular, a higher enhancement in β-glucosidase activity was obtained, which plays a key role in improving cellulase function. The DESs prolonged the half-life of cellulase. The addition of 10 % (v/v) ChCl/1,3-propanediol (1:2) and 5 % (v/v) Bet/glycerol (1:2) resulted in the most significant improvement, with the half-life of cellulase reaching three times the initial half-life value. The above both DESs allowed the cellulase to maintain an excellent level of activity over time, even after 72 h. The DESs tightened the structure of the enzyme by changing its secondary structure, which had a stabilizing effect on the overall conformation. The density functional theory calculations of DESs revealed that the DES with a more stable structure more effectively protected the active conformation. The molecular dynamics simulation on β-glucosidase indicated that the hydrogen bond donor end of the DES primarily formed hydrogen bonds with the enzyme, which induced a spatially compact conformation by increasing the α-helix content while reducing the β-sheet content. It also restricted the movement of amino acids, particularly stabilizing the active site, and increased the hydrophilic region, thereby enhancing enzyme activity and stability. In addition, the DES formed a protective layer that prevented the unproductive adsorption of lignin on the enzymes during the hydrolysis of lignocellulosic feedstocks. These multiple improvements provided by DESs significantly increased the enzymatic saccharification efficiency of corn stover and poplar. [ABSTRACT FROM AUTHOR]

The lignin-derived inhibition on cellulase is the most critical bottleneck that must be addressed when converting lignocellulosic biomass through the sugar-platform process. Herein, a novel modified cellulase with high activity, great stability and excellent lignin-resistant ability was synthesized by chemical modification using hydrophilic and negatively charged polyethylene glycol (PEG) derivatives including HO-PEG-COOH and HOOC-PEG-COOH. Both modifications obviously increased the cellulase activity, with 37% (HO-PEG-COOH modification) and 53% (HOOC-PEG-COOH modification) increase, respectively. In addition, the stabilities of both modified cellulases were significantly enhanced, seeing from their much higher activities at all selected temperatures and pH values. The modification resulted in a movement of tryptophan residues to the exterior of enzyme, allowing the substrate chains to be more easily recruited and thus initiating the hydrolysis. The compactness of secondary structure for enzyme was improved, especially, the α-helix content was increased which requires strong hydrogen bonding to generate it, thus enhancing the stability. High hydrophilicity, negative charge and compactness of enzyme conferred modified cellulase good ability to resist lignin adsorption and destruction, thus improving the enzymatic hydrolysis of corn stover. [Display omitted] • Cellulase activity is significantly improved by up to 52% after chemical modification. • The migration of Trp residues allows the hydrolysis to be more easily initiated. • High compactness gives modified cellulase great stability and low level of damage by lignin. • High hydrophilicity and negative charge of modified cellulase prevent lignin adsorption. • The novel modified cellulase exhibits promising potential in converting corn stover. [ABSTRACT FROM AUTHOR]