The Effects of Noncellulosic Compounds on the Nanoscale Interaction Forces Measured between Carbohydrate-Binding Module and Lignocellulosic Biomass

Abstract

The lack of fundamental understanding of the types of forces that govern how cellulose-degrading enzymes interact with cellulosic and noncellulosic components of lignocellulosic surfaces limits the design of new strategies for efficient conversion of biomass to bioethanol. In a step to improve our fundamental understanding of such interactions, nanoscale forces acting between a model cellulase-a carbohydrate-binding module (CBM) of cellobiohydrolase I (CBH I)-and a set of lignocellulosic substrates with controlled composition were measured using atomic force microscopy (AFM). The three model substrates investigated were kraft (KP), sulfite (SP), and organosolv (OPP) pulped substrates. These substrates varied in their surface lignin coverage, lignin type, and xylan and acetone extractives' content. Our results indicated that the overall adhesion forces of biomass to CBM increased linearly with surface lignin coverage with kraft lignin showing the highest forces among lignin types investigated. When the overall adhesion forces were decoupled into specific and nonspecific component forces via the Poisson statistical model, hydrophobic and Lifshitz-van der Waals (LW) forces dominated the binding forces of CBM to kraft lignin, whereas permanent dipole-dipole interactions and electrostatic forces facilitated the interactions of lignosulfonates to CBM. Xylan and acetone extractives' content increased the attractive forces between CBM and lignin-free substrates, most likely through hydrogen bonding forces. When the substrates treated differently were compared, it was found that both the differences in specific and nonspecific forces between lignin-containing and lignin-free substrates were the least for OPP. Therefore, cellulase enzymes represented by CBM would weakly bind to organosolv lignin. This will facilitate an easy enzyme recovery compared to other substrates treated with kraft or sulfite pulping. Our results also suggest that altering the surface hydrophobicity and the surface energy of lignin that facilitates the LW forces should be a priori to avoid nonproductive binding of cellulase to kraft lignin.

Acoustic Force Spectroscopy Reveals Subtle Differences in Cellulose Unbinding Behavior of Carbohydrate-Binding Modules

“…The fact that no rupture forces greater than 25 pN were measured for Y67A at 1 pN/s could be related to the difference in sample size (N=259 vs. N=138 for WT and Y67A, respectively) as the tail of larger rupture forces at 0.1 pN/s is similar for WT and Y67A (N=161 vs N=159 for WT and Y67A, respectively). A similar shape of rupture force distributions was observed in previous AFM-based studies for CBM3a (27) and CBM1 (24, 56) but previous AFM analysis showed a more Gaussian-like distribution for CBM3a (28), (25). King et al (27) showed that specific binding of CBM3a can be blocked with the addition of NCC and restored by washing the CBM-functionalized AFM tip with an excess of water.…”

NMR Analysis on Molecular Interaction of Lignin with Amino Acid Residues of Carbohydrate-Binding Module from Trichoderma reesei Cel7A

“…The exceptions are the studies of Guo and Arslan. They studied the affinities of Tr CBM1 to various cellulose substrates 25 as well as the binding behavior of Tr CBM1 to lignocellulosic substrates using an atomic force microscope 26 . These reports described the expression and purification of Tr CBM1.…”

Characterizing interaction forces between actin and proteins of the tropomodulin family reveals the presence of the N-terminal actin-binding site in leiomodin

“…AFM provides a valuable approach to quantify interactions between biomolecules [52]. AFM’s ability to quantify biomolecular forces with pico-newton resolution in liquid environments [47, 57] allowed researchers to investigate protein-ligand [58], protein-DNA [59], protein-protein [60], and protein-biomolecule [61] interactions at the nanoscale. The ability to work with full-length proteins, the use of relatively low protein concentrations (~0.5 to 3 μM in this study) and stability of actin in a monomeric form make AFM a promising technique for measuring forces of interaction of G-actin with other actin-nucleating and actin-sequestering proteins.…”