Neutron Reflectometry and QCM-D Study of the Interaction of Cellulases with Films of Amorphous Cellulose

Abstract

Improving the efficiency of enzymatic hydrolysis of cellulose is one of the key technological hurdles to reduce the cost of producing ethanol and other transportation fuels from lignocellulosic material. A better understanding of how soluble enzymes interact with insoluble cellulose will aid in the design of more efficient enzyme systems. We report a study involving neutron reflectometry (NR) and quartz crystal microbalance with dissipation monitoring (QCM-D) of the interaction of a fungal enzyme extract ( T. viride ) and an endoglucanse from A. niger with amorphous cellulose films. The use of amorphous cellulose is motivated by that the fact that several biomass pretreatments currently under investigation disrupt the native crystalline structure of cellulose and increase the amorphous content. NR reveals the profile of water through the film at nanometer resolution and is highly sensitive to interfacial roughness, whereas QCM-D provides changes in mass and film stiffness. NR can be performed using either H(2)O- or D(2)O-based aqueous reservoirs. NR measurement of swelling of a cellulose film in D(2)O and in H(2)O revealed that D/H exchange on the cellulose chains must be taken into account when a D(2)O-based reservoir is used. The results also show that cellulose films swell slightly more in D(2)O than in H(2)O. Regarding enzymatic digestion, at 20 °C in H(2)O buffer the T. viride cocktail rapidly digested the entire film, initially roughening the surface, followed by penetration and activity throughout the bulk of the film. In contrast, over the same time period, the endoglucanase was active mainly at the surface of the film and did not increase the surface roughness.

Preferential Adsorption and Activity of Monocomponent Cellulases on Lignocellulose Thin Films with Varying Lignin Content

“…This strong increase in dissipation indicates penetration of the enzyme into and digestion within the bulk of the film, causing water swelling and decreasing the stiffness throughout the bulk of the film. 22,25,26 If the hydrolysis continues, the dissipation value will reach a maximum at a certain point after which it starts to decrease, because the film becomes denser, and the surface turns into cellulose clusters, which contain the less hydrolyzable residual cellulose chains. 20,22,23,25,26 However, in the current study, the buffer rinsing of the film was carried out before reaching this point, so the results in Figure 2 correspond to the early stages of substrate hydrolysis.…”

“…As mentioned above, this phase is associated with the penetration of the enzyme into and digestion within the bulk of the film. 25,26 In fact, it has been proposed that the enzyme penetrates by the CBD hopping from one cellulose binding site to another. 26 Therefore, there is indication that the CBH I core is not able to penetrate, and its action is localized only on the surface of film.…”

“…The pump was then stopped and started again (0.1 mL/min) after 3 h (4 h in the case of EG I) of treatment to rinse the system with buffer solution. All measurements were recorded at 5 MHz fundamental resonance frequency and its overtones corresponding to 15,25,35,55, and 75 MHz. The third overtone (15 MHz) was used for data processing.…”

Tailor-made functional surfaces based on cellulose-derived materials

“…Smooth and uniform cellulose surfaces have been prepared via a variety of approaches, including (1) amorphous cellulose surfaces regenerated from trimethylsilyl cellulose (TMSC) thin films Cheng et al 2011;Kontturi et al 2003;Schaub et al 1993), (2) cellulose II surfaces regenerated from the solutions of cellulose in dimethylacetamide/lithium chloride (DMAc/LiCl) (Aulin et al 2009;Eriksson et al 2005) (Maurer et al 2012;Notley and Wågberg 2005), (3) nanocrystalline cellulose surfaces spin-coated from the aqueous colloidal suspensions of acid-hydrolyzed cellulose nanocrystals (or nanowhiskers) (Edgar and Gray 2003;Kittle et al 2012), and (4) nanofibrillar cellulose surfaces spincoated from aqueous dispersions of micro-or nanofibrillar cellulose (Ahola et al 2008a;Ahola et al 2008b). Those well-defined cellulose surfaces provide simplified and convenient platforms to study their swelling, enzymatic degradation, surface modification, and interactions with polymers, surfactants, polysaccharides, and proteins under carefully controlled conditions.…”

Adsorption of cationic polyacrylamide at the cellulose–liquid interface: A neutron reflectometry study

“…Optimal fitting of the NR results with the model was achieved by assuming the proton-deuterium exchange [33] in the hydroxyl groups of the cellulose in D 2 O [34] which corresponds to a cellulose density of 1.48 g/cm 3 . As the regenerated cellulose films are completely amorphous, we assumed full exchange would occur.…”