Measurements of single molecular affinity interactions between carbohydrate-binding modules and crystalline cellulose fibrils

Abstract

Combining atomic force microscopy (AFM) recognition imaging and single molecule dynamic force spectroscopy (SMDFS), we studied the single molecule affinity interactions between the carbohydrate-binding module (CBM) and plant cell wall cellulose using the CBM3a (from Clostridium thermocellum) and CBM2a (from Cellvibrio japonicus) functionalized AFM tips. The binding efficiencies of the CBMs to the cellulose were determined by the binding areas on the crystalline cellulose fibrils surface using the recognition imaging. Several dynamic and kinetic parameters, such as the reconstructed free energy change, energy barrier and bond lifetime constant, were also obtained based on the measured single molecule unbinding forces, which are used to illuminate the affinity of the CBMs binding to the natural and single cellulose surface from a totally different aspect. It was found that CBM3a has a little higher binding efficiency and affinity than CBM2a to both natural and extracted cellulose surfaces and both the CBMs have higher affinities to the natural cell wall cellulose compared to the extracted single cellulose. The in-depth understanding of the binding mechanisms of the CBM-cellulose interactions of this study may pave the way for more efficient plant cell wall degradation and eventually facilitate biofuel production.

The cohesin module is a major determinant of cellulosome mechanical stability

“…On the other hand, the mechanical stability of the CBM-cellulose interaction has also been experimentally addressed (35) and found relatively low unbinding forces. Nevertheless, since multiple binding sites are available on the surface of cellulose, it is reasonable to expect that the cellulosome would be effectively anchored to the cellulose for extended periods of time, thus exposing cohesins to forces.…”

Nanometrology of Biomass for Bioenergy: The Role of Atomic Force Microscopy and Spectroscopy in Plant Cell Characterization

“…AM-AFM has been successful in characterizing biomass (Kirby et al, 1996;Salvadori et al, 2014;Farahi et al, 2017) and in particular the architecture of plant cell walls at the molecular level which in turn is essential in improving lignocellulosic feedstock properties (Zhang et al, 2013b(Zhang et al, , 2017Keplinger et al, 2014;Torode et al, 2018). AM-AFM mapping of cellulose topography revealed the right-handed twisted nature of microfibrils and the periodic distribution of glucose and fiber unit along the microfibrils (Hanley et al, 1997).…”

Mapping out the structural changes of natural and pretreated plant cell wall surfaces by atomic force microscopy single molecular recognition imaging

“…The results indicated a pronounced, specific unbinding force peak in the force histograms for CBM3a-crystalline cellulose interaction, while near-zero, non-specific force was obtained when taking the force data for all the non- cellulose areas on the poplar slice surface. For bare AFM tip, only the near-zero, non-specific force was observed for all the areas on the poplar slice surface [36,37]. In this work, we also characterized the natural cell wall of switchgrass and corn stover and the representative topography and recognition images are shown in Figure 2.…”

“…All images were taken using non-contact, top magnetic AC (TopMAC) mode under PicoTREC (Agilent Technologies, Santa Clara, CA). The topography (height) and recognition images were conducted simultaneously [32,37]. The 200–500 μm hand-cut, natural pieces of each biomass sample after mesh screening (200–500 μm in size) before pretreatments and pure microcrystalline cellulose Avicel PH-105 (FMC BioPolymer, Philadelphia, PA, nominal particle size: 20 μm) were also imaged.…”

“…The AFM recognition imaging is also used as a reliable and efficient tool in studying carbohydrate-protein interactions [35]. Recently, AFM recognition imaging and SMDFS have been applied to map the natural and pretreated poplar cell wall surface and study the affinity between a non-catalytic family 3 carbohydrate-binding module (CBM3a) and crystalline cellulose [36,37]. The crystalline cellulose exposed on cell wall surface was specifically recognized and its surface coverage was quantified [37].…”