Structural insights into the affinity of Cel7A carbohydrate-binding module for lignin

Area of Science:

• Biotechnology
• Biochemistry
• Renewable Energy

Background:

• High costs of hydrolytic enzymes hinder lignocellulosic biofuel production.
• Non-productive adsorption of enzymes to lignin increases required enzyme loadings.
• Limited research exists on engineering enzymes to reduce lignin binding.

Purpose of the Study:

• To elucidate the structural basis of lignin affinity in Trichoderma reesei Cel7A carbohydrate binding module (CBM).
• To engineer CBM mutants with reduced lignin binding while maintaining cellulose affinity.

Main Methods:

• Alanine-scanning mutagenesis was used to create T. reesei Cel7A CBM mutants.
• Mutants were combined with a Talaromyces emersonii Cel7A catalytic domain.
• Binding affinities to cellulose and lignin were screened for 31 mutants.

Main Results:

• Mutation of aromatic and polar residues on the CBM planar face reduced binding to both cellulose and lignin.
• Cellulose and lignin affinities were highly correlated, but some mutants showed selective affinity reduction.
• Four mutants with increased cellulose selectivity did not improve cellulose hydrolysis in the presence of lignin.

Conclusions:

• The cellulose-binding face of the CBM is also responsible for lignin affinity.
• Reducing lignin affinity while maintaining cellulose affinity is crucial for improving enzyme efficacy.
• This study provides insights for engineering family one CBMs for enhanced biofuel production.