Comparative Evaluation of Adsorption of Major Enzymes in a Cellulase Cocktail Obtained from Trichoderma reesei onto Different Types of Lignin

Abstract

Cellulase adsorption onto lignin decreases the productivity of enzymatic hydrolysis of lignocellulosic biomass. Here, adsorption of enzymes onto different types of lignin was investigated, and the five major enzymes—cellobiohydrolases (CBHs), endoglucanase (Cel7B), β-glucosidase (Cel3A), xylanase (XYNIV), and mannanase (Man5A)—in a cellulase cocktail obtained from Trichoderma reesei were individually analyzed through SDS-PAGE and zymogram assay. Lignin was isolated from woody (oak and pine lignin) and herbaceous (rice straw and kenaf lignin) plants. The relative adsorption of CBHs compared to the control was in the range of 14.15–18.61%. The carbohydrate binding motif (CBM) of the CBHs contributed to higher adsorption levels in oak and kenaf lignin, compared to those in pine and rice lignin. The adsorption of endoglucanase (Cel7B) by herbaceous plant lignin was two times higher than that of woody lignin, whereas XYNIV showed the opposite pattern. β-glucosidase (Cel3A) displayed the highest and lowest adsorption ratios on rice straw and kenaf lignin, respectively. Mannanase (Man5A) was found to have the lowest adsorption ratio on pine lignin. Our results showed that the hydrophobic properties of CBM and the enzyme structures are key factors in adsorption onto lignin, whereas the properties of specific lignin types indirectly affect adsorption.

Implementation of CDH in LPMO-Containing Minimal Cellulase Cocktails to Tune Bioproduct Production from Lignocellulosic Biomass

“…Notably, the lignin remaining after the pretreatment may contribute to redox side reactions that consume or produce H 2 O 2 , and it may also prevent LPMOs from optimally accessing cellulose. Indeed, it has been shown that the hydrophobic properties of CBM1 (which is present in both Pa AA9E and Pa AA9H) contribute to the nonproductive binding to lignin. , The biomass used in this study indeed contains high concentrations of remaining lignins (Table S3), which could explain the lack of a boosting effect coming from the addition of the tested LPMOs. Therefore, although Pa AA9E and Pa AA9H are efficient cellulolytic AA9s, it appears clearly that observing a high activity on somehow simple cellulosic substrates (such as PASC or Avicel) cannot be taken as a token of efficiency once mixed with cellulases on more complex biomass.…”

Fluorescent Imaging of Extracellular Fungal Enzymes Bound onto Plant Cell Walls

“…It is worth noting that protein adsorption on solid substrates is a dynamic process that involves continuous structural changes in proteins as they reversibly adsorb and desorb. However, as the process progresses, numerous rearrangements between the two states may compromise the protein's structural integrity, resulting in irreversible structural alterations and protein activity loss [47,48]. For most of the enzymes, the highest fluorescence intensity was localized on the inner part of the cell wall, corresponding to secondary cell wall layers (SCW) S3 and S2 sublayers.…”