Method for characterizing interaction force between lignin and cellulase

A method for characterizing an interaction force between a lignin model substance and cellulase, comprising the following steps:

• (1) a preparation of a lignin model substance film:
• preparing a 0.5% lignin model substance solution, then dropwise adding 100 μL of the 0.5% lignin model substance solution on a silicon wafer using a flat mouth syringe, after the 0.5% lignin model substance solution diffuses to an entire silicon wafer surface, allowing the silicon wafer to stand for 1 min, and using a spin coater to spin-coat at a speed of 5000 r/min for 1 min, with 3 repeats;
• vacuum-drying a prepared lignin film at 40° C. for 4 h, and then soaking a dried lignin film in deionized water for 1 day;
• replacing the deionized water every 2 h to remove dimethylsulfoxide (DMSO);
• vacuum-drying a soaked film at 40° C. for 12 h;
• wherein a lignin model substance is one or more of the following substances:
• (2) an atomic force microscopy (AFM) probe modification:
• modifying an AFM probe coated with a gold film on a surface with HS-PEG-COOH and then activating a carboxyl group, immersing the AFM probe after a carboxyl group activation in a phosphate buffer saline (PBS) solution of the cellulase for an incubation, and after the AFM probe modification is completed, placing a washed AFM probe in a PBS buffer solution for a storage and a later use;
• (3) using an AFM instrument to test a lignin model substance-cellulase interaction force in a liquid phase environment:
• detecting a dissociation rate of the interaction between the lignin model substance molecules modified on the film and the cellulase protein modified on the tip of the AFM probe, evaluating a binding ability of the lignin model substance and the cellulase by a dissociation rate constant of an interaction reaction, and after counting a number of representative force-displacement curves at each loading rate, calculating a dissociation force between the lignin model substance and the cellulase using the following Bell-Evans equation:
• F
• =
• k
• B
• ⁢
• T
• x
• β
• ×
• ln
• ⁢
• Rx
• β
• k
• B
• ⁢
• Tk
• off
• wherein F represents the dissociation force, Xβ is a distance from a lowest energy point of a binding state to a molecular activation state on a dissociation path, R is a dissociation force loading rate, R=keff×v, where keff is an effective elastic coefficient of an AFM probe cantilever, and v is a retraction rate of the AFM probe, koff is a dissociation rate constant when the lignin model substance-cellulase interaction force is zero, T is a thermodynamic temperature, and kB is a Boltzmann constant.